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How to Write a Great Hypothesis

Hypothesis Definition, Format, Examples, and Tips

Kendra Cherry, MS, is a psychosocial rehabilitation specialist, psychology educator, and author of the "Everything Psychology Book."

Amy Morin, LCSW, is a psychotherapist and international bestselling author. Her books, including "13 Things Mentally Strong People Don't Do," have been translated into more than 40 languages. Her TEDx talk,  "The Secret of Becoming Mentally Strong," is one of the most viewed talks of all time.

Verywell / Alex Dos Diaz

• The Scientific Method

Hypothesis Format

Falsifiability of a hypothesis.

• Operationalization

Hypothesis Types

Hypotheses examples.

• Collecting Data

A hypothesis is a tentative statement about the relationship between two or more variables. It is a specific, testable prediction about what you expect to happen in a study. It is a preliminary answer to your question that helps guide the research process.

Consider a study designed to examine the relationship between sleep deprivation and test performance. The hypothesis might be: "This study is designed to assess the hypothesis that sleep-deprived people will perform worse on a test than individuals who are not sleep-deprived."

At a Glance

A hypothesis is crucial to scientific research because it offers a clear direction for what the researchers are looking to find. This allows them to design experiments to test their predictions and add to our scientific knowledge about the world. This article explores how a hypothesis is used in psychology research, how to write a good hypothesis, and the different types of hypotheses you might use.

The Hypothesis in the Scientific Method

In the scientific method , whether it involves research in psychology, biology, or some other area, a hypothesis represents what the researchers think will happen in an experiment. The scientific method involves the following steps:

• Forming a question
• Performing background research
• Creating a hypothesis
• Designing an experiment
• Collecting data
• Analyzing the results
• Drawing conclusions
• Communicating the results

The hypothesis is a prediction, but it involves more than a guess. Most of the time, the hypothesis begins with a question which is then explored through background research. At this point, researchers then begin to develop a testable hypothesis.

Unless you are creating an exploratory study, your hypothesis should always explain what you  expect  to happen.

In a study exploring the effects of a particular drug, the hypothesis might be that researchers expect the drug to have some type of effect on the symptoms of a specific illness. In psychology, the hypothesis might focus on how a certain aspect of the environment might influence a particular behavior.

Remember, a hypothesis does not have to be correct. While the hypothesis predicts what the researchers expect to see, the goal of the research is to determine whether this guess is right or wrong. When conducting an experiment, researchers might explore numerous factors to determine which ones might contribute to the ultimate outcome.

In many cases, researchers may find that the results of an experiment  do not  support the original hypothesis. When writing up these results, the researchers might suggest other options that should be explored in future studies.

In many cases, researchers might draw a hypothesis from a specific theory or build on previous research. For example, prior research has shown that stress can impact the immune system. So a researcher might hypothesize: "People with high-stress levels will be more likely to contract a common cold after being exposed to the virus than people who have low-stress levels."

In other instances, researchers might look at commonly held beliefs or folk wisdom. "Birds of a feather flock together" is one example of folk adage that a psychologist might try to investigate. The researcher might pose a specific hypothesis that "People tend to select romantic partners who are similar to them in interests and educational level."

Elements of a Good Hypothesis

So how do you write a good hypothesis? When trying to come up with a hypothesis for your research or experiments, ask yourself the following questions:

• Is your hypothesis based on your research on a topic?
• Can your hypothesis be tested?
• Does your hypothesis include independent and dependent variables?

Before you come up with a specific hypothesis, spend some time doing background research. Once you have completed a literature review, start thinking about potential questions you still have. Pay attention to the discussion section in the  journal articles you read . Many authors will suggest questions that still need to be explored.

How to Formulate a Good Hypothesis

To form a hypothesis, you should take these steps:

• Collect as many observations about a topic or problem as you can.
• Evaluate these observations and look for possible causes of the problem.
• Create a list of possible explanations that you might want to explore.
• After you have developed some possible hypotheses, think of ways that you could confirm or disprove each hypothesis through experimentation. This is known as falsifiability.

In the scientific method ,  falsifiability is an important part of any valid hypothesis. In order to test a claim scientifically, it must be possible that the claim could be proven false.

Students sometimes confuse the idea of falsifiability with the idea that it means that something is false, which is not the case. What falsifiability means is that  if  something was false, then it is possible to demonstrate that it is false.

One of the hallmarks of pseudoscience is that it makes claims that cannot be refuted or proven false.

The Importance of Operational Definitions

A variable is a factor or element that can be changed and manipulated in ways that are observable and measurable. However, the researcher must also define how the variable will be manipulated and measured in the study.

Operational definitions are specific definitions for all relevant factors in a study. This process helps make vague or ambiguous concepts detailed and measurable.

For example, a researcher might operationally define the variable " test anxiety " as the results of a self-report measure of anxiety experienced during an exam. A "study habits" variable might be defined by the amount of studying that actually occurs as measured by time.

These precise descriptions are important because many things can be measured in various ways. Clearly defining these variables and how they are measured helps ensure that other researchers can replicate your results.

Replicability

One of the basic principles of any type of scientific research is that the results must be replicable.

Replication means repeating an experiment in the same way to produce the same results. By clearly detailing the specifics of how the variables were measured and manipulated, other researchers can better understand the results and repeat the study if needed.

Some variables are more difficult than others to define. For example, how would you operationally define a variable such as aggression ? For obvious ethical reasons, researchers cannot create a situation in which a person behaves aggressively toward others.

To measure this variable, the researcher must devise a measurement that assesses aggressive behavior without harming others. The researcher might utilize a simulated task to measure aggressiveness in this situation.

Hypothesis Checklist

• Does your hypothesis focus on something that you can actually test?
• Does your hypothesis include both an independent and dependent variable?
• Can you manipulate the variables?
• Can your hypothesis be tested without violating ethical standards?

The hypothesis you use will depend on what you are investigating and hoping to find. Some of the main types of hypotheses that you might use include:

• Simple hypothesis : This type of hypothesis suggests there is a relationship between one independent variable and one dependent variable.
• Complex hypothesis : This type suggests a relationship between three or more variables, such as two independent and dependent variables.
• Null hypothesis : This hypothesis suggests no relationship exists between two or more variables.
• Alternative hypothesis : This hypothesis states the opposite of the null hypothesis.
• Statistical hypothesis : This hypothesis uses statistical analysis to evaluate a representative population sample and then generalizes the findings to the larger group.
• Logical hypothesis : This hypothesis assumes a relationship between variables without collecting data or evidence.

A hypothesis often follows a basic format of "If {this happens} then {this will happen}." One way to structure your hypothesis is to describe what will happen to the  dependent variable  if you change the  independent variable .

The basic format might be: "If {these changes are made to a certain independent variable}, then we will observe {a change in a specific dependent variable}."

A few examples of simple hypotheses:

• "Students who eat breakfast will perform better on a math exam than students who do not eat breakfast."
• "Students who experience test anxiety before an English exam will get lower scores than students who do not experience test anxiety."​
• "Motorists who talk on the phone while driving will be more likely to make errors on a driving course than those who do not talk on the phone."
• "Children who receive a new reading intervention will have higher reading scores than students who do not receive the intervention."

Examples of a complex hypothesis include:

• "People with high-sugar diets and sedentary activity levels are more likely to develop depression."
• "Younger people who are regularly exposed to green, outdoor areas have better subjective well-being than older adults who have limited exposure to green spaces."

Examples of a null hypothesis include:

• "There is no difference in anxiety levels between people who take St. John's wort supplements and those who do not."
• "There is no difference in scores on a memory recall task between children and adults."
• "There is no difference in aggression levels between children who play first-person shooter games and those who do not."

Examples of an alternative hypothesis:

• "People who take St. John's wort supplements will have less anxiety than those who do not."
• "Adults will perform better on a memory task than children."
• "Children who play first-person shooter games will show higher levels of aggression than children who do not." 

Collecting Data on Your Hypothesis

Once a researcher has formed a testable hypothesis, the next step is to select a research design and start collecting data. The research method depends largely on exactly what they are studying. There are two basic types of research methods: descriptive research and experimental research.

Descriptive Research Methods

Descriptive research such as  case studies ,  naturalistic observations , and surveys are often used when  conducting an experiment is difficult or impossible. These methods are best used to describe different aspects of a behavior or psychological phenomenon.

Once a researcher has collected data using descriptive methods, a  correlational study  can examine how the variables are related. This research method might be used to investigate a hypothesis that is difficult to test experimentally.

Experimental Research Methods

Experimental methods  are used to demonstrate causal relationships between variables. In an experiment, the researcher systematically manipulates a variable of interest (known as the independent variable) and measures the effect on another variable (known as the dependent variable).

Unlike correlational studies, which can only be used to determine if there is a relationship between two variables, experimental methods can be used to determine the actual nature of the relationship—whether changes in one variable actually  cause  another to change.

The hypothesis is a critical part of any scientific exploration. It represents what researchers expect to find in a study or experiment. In situations where the hypothesis is unsupported by the research, the research still has value. Such research helps us better understand how different aspects of the natural world relate to one another. It also helps us develop new hypotheses that can then be tested in the future.

Thompson WH, Skau S. On the scope of scientific hypotheses .  R Soc Open Sci . 2023;10(8):230607. doi:10.1098/rsos.230607

Taran S, Adhikari NKJ, Fan E. Falsifiability in medicine: what clinicians can learn from Karl Popper [published correction appears in Intensive Care Med. 2021 Jun 17;:].  Intensive Care Med . 2021;47(9):1054-1056. doi:10.1007/s00134-021-06432-z

Eyler AA. Research Methods for Public Health . 1st ed. Springer Publishing Company; 2020. doi:10.1891/9780826182067.0004

Nosek BA, Errington TM. What is replication ?  PLoS Biol . 2020;18(3):e3000691. doi:10.1371/journal.pbio.3000691

Aggarwal R, Ranganathan P. Study designs: Part 2 - Descriptive studies .  Perspect Clin Res . 2019;10(1):34-36. doi:10.4103/picr.PICR_154_18

Nevid J. Psychology: Concepts and Applications. Wadworth, 2013.

By Kendra Cherry, MSEd Kendra Cherry, MS, is a psychosocial rehabilitation specialist, psychology educator, and author of the "Everything Psychology Book."

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Home » What is a Hypothesis – Types, Examples and Writing Guide

What is a Hypothesis – Types, Examples and Writing Guide

Table of Contents

Definition:

Hypothesis is an educated guess or proposed explanation for a phenomenon, based on some initial observations or data. It is a tentative statement that can be tested and potentially proven or disproven through further investigation and experimentation.

Hypothesis is often used in scientific research to guide the design of experiments and the collection and analysis of data. It is an essential element of the scientific method, as it allows researchers to make predictions about the outcome of their experiments and to test those predictions to determine their accuracy.

Types of Hypothesis

Types of Hypothesis are as follows:

Research Hypothesis

A research hypothesis is a statement that predicts a relationship between variables. It is usually formulated as a specific statement that can be tested through research, and it is often used in scientific research to guide the design of experiments.

Null Hypothesis

The null hypothesis is a statement that assumes there is no significant difference or relationship between variables. It is often used as a starting point for testing the research hypothesis, and if the results of the study reject the null hypothesis, it suggests that there is a significant difference or relationship between variables.

Alternative Hypothesis

An alternative hypothesis is a statement that assumes there is a significant difference or relationship between variables. It is often used as an alternative to the null hypothesis and is tested against the null hypothesis to determine which statement is more accurate.

Directional Hypothesis

A directional hypothesis is a statement that predicts the direction of the relationship between variables. For example, a researcher might predict that increasing the amount of exercise will result in a decrease in body weight.

Non-directional Hypothesis

A non-directional hypothesis is a statement that predicts the relationship between variables but does not specify the direction. For example, a researcher might predict that there is a relationship between the amount of exercise and body weight, but they do not specify whether increasing or decreasing exercise will affect body weight.

Statistical Hypothesis

A statistical hypothesis is a statement that assumes a particular statistical model or distribution for the data. It is often used in statistical analysis to test the significance of a particular result.

Composite Hypothesis

A composite hypothesis is a statement that assumes more than one condition or outcome. It can be divided into several sub-hypotheses, each of which represents a different possible outcome.

Empirical Hypothesis

An empirical hypothesis is a statement that is based on observed phenomena or data. It is often used in scientific research to develop theories or models that explain the observed phenomena.

Simple Hypothesis

A simple hypothesis is a statement that assumes only one outcome or condition. It is often used in scientific research to test a single variable or factor.

Complex Hypothesis

A complex hypothesis is a statement that assumes multiple outcomes or conditions. It is often used in scientific research to test the effects of multiple variables or factors on a particular outcome.

Applications of Hypothesis

Hypotheses are used in various fields to guide research and make predictions about the outcomes of experiments or observations. Here are some examples of how hypotheses are applied in different fields:

• Science : In scientific research, hypotheses are used to test the validity of theories and models that explain natural phenomena. For example, a hypothesis might be formulated to test the effects of a particular variable on a natural system, such as the effects of climate change on an ecosystem.
• Medicine : In medical research, hypotheses are used to test the effectiveness of treatments and therapies for specific conditions. For example, a hypothesis might be formulated to test the effects of a new drug on a particular disease.
• Psychology : In psychology, hypotheses are used to test theories and models of human behavior and cognition. For example, a hypothesis might be formulated to test the effects of a particular stimulus on the brain or behavior.
• Sociology : In sociology, hypotheses are used to test theories and models of social phenomena, such as the effects of social structures or institutions on human behavior. For example, a hypothesis might be formulated to test the effects of income inequality on crime rates.
• Business : In business research, hypotheses are used to test the validity of theories and models that explain business phenomena, such as consumer behavior or market trends. For example, a hypothesis might be formulated to test the effects of a new marketing campaign on consumer buying behavior.
• Engineering : In engineering, hypotheses are used to test the effectiveness of new technologies or designs. For example, a hypothesis might be formulated to test the efficiency of a new solar panel design.

How to write a Hypothesis

Here are the steps to follow when writing a hypothesis:

Identify the Research Question

The first step is to identify the research question that you want to answer through your study. This question should be clear, specific, and focused. It should be something that can be investigated empirically and that has some relevance or significance in the field.

Conduct a Literature Review

Before writing your hypothesis, it’s essential to conduct a thorough literature review to understand what is already known about the topic. This will help you to identify the research gap and formulate a hypothesis that builds on existing knowledge.

Determine the Variables

The next step is to identify the variables involved in the research question. A variable is any characteristic or factor that can vary or change. There are two types of variables: independent and dependent. The independent variable is the one that is manipulated or changed by the researcher, while the dependent variable is the one that is measured or observed as a result of the independent variable.

Formulate the Hypothesis

Based on the research question and the variables involved, you can now formulate your hypothesis. A hypothesis should be a clear and concise statement that predicts the relationship between the variables. It should be testable through empirical research and based on existing theory or evidence.

Write the Null Hypothesis

The null hypothesis is the opposite of the alternative hypothesis, which is the hypothesis that you are testing. The null hypothesis states that there is no significant difference or relationship between the variables. It is important to write the null hypothesis because it allows you to compare your results with what would be expected by chance.

Refine the Hypothesis

After formulating the hypothesis, it’s important to refine it and make it more precise. This may involve clarifying the variables, specifying the direction of the relationship, or making the hypothesis more testable.

Examples of Hypothesis

Here are a few examples of hypotheses in different fields:

• Psychology : “Increased exposure to violent video games leads to increased aggressive behavior in adolescents.”
• Biology : “Higher levels of carbon dioxide in the atmosphere will lead to increased plant growth.”
• Sociology : “Individuals who grow up in households with higher socioeconomic status will have higher levels of education and income as adults.”
• Education : “Implementing a new teaching method will result in higher student achievement scores.”
• Marketing : “Customers who receive a personalized email will be more likely to make a purchase than those who receive a generic email.”
• Physics : “An increase in temperature will cause an increase in the volume of a gas, assuming all other variables remain constant.”
• Medicine : “Consuming a diet high in saturated fats will increase the risk of developing heart disease.”

Purpose of Hypothesis

The purpose of a hypothesis is to provide a testable explanation for an observed phenomenon or a prediction of a future outcome based on existing knowledge or theories. A hypothesis is an essential part of the scientific method and helps to guide the research process by providing a clear focus for investigation. It enables scientists to design experiments or studies to gather evidence and data that can support or refute the proposed explanation or prediction.

The formulation of a hypothesis is based on existing knowledge, observations, and theories, and it should be specific, testable, and falsifiable. A specific hypothesis helps to define the research question, which is important in the research process as it guides the selection of an appropriate research design and methodology. Testability of the hypothesis means that it can be proven or disproven through empirical data collection and analysis. Falsifiability means that the hypothesis should be formulated in such a way that it can be proven wrong if it is incorrect.

In addition to guiding the research process, the testing of hypotheses can lead to new discoveries and advancements in scientific knowledge. When a hypothesis is supported by the data, it can be used to develop new theories or models to explain the observed phenomenon. When a hypothesis is not supported by the data, it can help to refine existing theories or prompt the development of new hypotheses to explain the phenomenon.

When to use Hypothesis

Here are some common situations in which hypotheses are used:

• In scientific research , hypotheses are used to guide the design of experiments and to help researchers make predictions about the outcomes of those experiments.
• In social science research , hypotheses are used to test theories about human behavior, social relationships, and other phenomena.
• I n business , hypotheses can be used to guide decisions about marketing, product development, and other areas. For example, a hypothesis might be that a new product will sell well in a particular market, and this hypothesis can be tested through market research.

Characteristics of Hypothesis

Here are some common characteristics of a hypothesis:

• Testable : A hypothesis must be able to be tested through observation or experimentation. This means that it must be possible to collect data that will either support or refute the hypothesis.
• Falsifiable : A hypothesis must be able to be proven false if it is not supported by the data. If a hypothesis cannot be falsified, then it is not a scientific hypothesis.
• Clear and concise : A hypothesis should be stated in a clear and concise manner so that it can be easily understood and tested.
• Based on existing knowledge : A hypothesis should be based on existing knowledge and research in the field. It should not be based on personal beliefs or opinions.
• Specific : A hypothesis should be specific in terms of the variables being tested and the predicted outcome. This will help to ensure that the research is focused and well-designed.
• Tentative: A hypothesis is a tentative statement or assumption that requires further testing and evidence to be confirmed or refuted. It is not a final conclusion or assertion.
• Relevant : A hypothesis should be relevant to the research question or problem being studied. It should address a gap in knowledge or provide a new perspective on the issue.

Advantages of Hypothesis

Hypotheses have several advantages in scientific research and experimentation:

• Guides research: A hypothesis provides a clear and specific direction for research. It helps to focus the research question, select appropriate methods and variables, and interpret the results.
• Predictive powe r: A hypothesis makes predictions about the outcome of research, which can be tested through experimentation. This allows researchers to evaluate the validity of the hypothesis and make new discoveries.
• Facilitates communication: A hypothesis provides a common language and framework for scientists to communicate with one another about their research. This helps to facilitate the exchange of ideas and promotes collaboration.
• Efficient use of resources: A hypothesis helps researchers to use their time, resources, and funding efficiently by directing them towards specific research questions and methods that are most likely to yield results.
• Provides a basis for further research: A hypothesis that is supported by data provides a basis for further research and exploration. It can lead to new hypotheses, theories, and discoveries.
• Increases objectivity: A hypothesis can help to increase objectivity in research by providing a clear and specific framework for testing and interpreting results. This can reduce bias and increase the reliability of research findings.

Limitations of Hypothesis

Some Limitations of the Hypothesis are as follows:

• Limited to observable phenomena: Hypotheses are limited to observable phenomena and cannot account for unobservable or intangible factors. This means that some research questions may not be amenable to hypothesis testing.
• May be inaccurate or incomplete: Hypotheses are based on existing knowledge and research, which may be incomplete or inaccurate. This can lead to flawed hypotheses and erroneous conclusions.
• May be biased: Hypotheses may be biased by the researcher’s own beliefs, values, or assumptions. This can lead to selective interpretation of data and a lack of objectivity in research.
• Cannot prove causation: A hypothesis can only show a correlation between variables, but it cannot prove causation. This requires further experimentation and analysis.
• Limited to specific contexts: Hypotheses are limited to specific contexts and may not be generalizable to other situations or populations. This means that results may not be applicable in other contexts or may require further testing.
• May be affected by chance : Hypotheses may be affected by chance or random variation, which can obscure or distort the true relationship between variables.

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Research Hypothesis In Psychology: Types, & Examples

Saul Mcleod, PhD

Editor-in-Chief for Simply Psychology

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Saul Mcleod, PhD., is a qualified psychology teacher with over 18 years of experience in further and higher education. He has been published in peer-reviewed journals, including the Journal of Clinical Psychology.

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On This Page:

A research hypothesis, in its plural form “hypotheses,” is a specific, testable prediction about the anticipated results of a study, established at its outset. It is a key component of the scientific method .

Hypotheses connect theory to data and guide the research process towards expanding scientific understanding

Some key points about hypotheses:

• A hypothesis expresses an expected pattern or relationship. It connects the variables under investigation.
• It is stated in clear, precise terms before any data collection or analysis occurs. This makes the hypothesis testable.
• A hypothesis must be falsifiable. It should be possible, even if unlikely in practice, to collect data that disconfirms rather than supports the hypothesis.
• Hypotheses guide research. Scientists design studies to explicitly evaluate hypotheses about how nature works.
• For a hypothesis to be valid, it must be testable against empirical evidence. The evidence can then confirm or disprove the testable predictions.
• Hypotheses are informed by background knowledge and observation, but go beyond what is already known to propose an explanation of how or why something occurs.

Predictions typically arise from a thorough knowledge of the research literature, curiosity about real-world problems or implications, and integrating this to advance theory. They build on existing literature while providing new insight.

Types of Research Hypotheses

Alternative hypothesis.

The research hypothesis is often called the alternative or experimental hypothesis in experimental research.

It typically suggests a potential relationship between two key variables: the independent variable, which the researcher manipulates, and the dependent variable, which is measured based on those changes.

The alternative hypothesis states a relationship exists between the two variables being studied (one variable affects the other).

A hypothesis is a testable statement or prediction about the relationship between two or more variables. It is a key component of the scientific method. Some key points about hypotheses:

• Important hypotheses lead to predictions that can be tested empirically. The evidence can then confirm or disprove the testable predictions.

In summary, a hypothesis is a precise, testable statement of what researchers expect to happen in a study and why. Hypotheses connect theory to data and guide the research process towards expanding scientific understanding.

An experimental hypothesis predicts what change(s) will occur in the dependent variable when the independent variable is manipulated.

It states that the results are not due to chance and are significant in supporting the theory being investigated.

The alternative hypothesis can be directional, indicating a specific direction of the effect, or non-directional, suggesting a difference without specifying its nature. It’s what researchers aim to support or demonstrate through their study.

Null Hypothesis

The null hypothesis states no relationship exists between the two variables being studied (one variable does not affect the other). There will be no changes in the dependent variable due to manipulating the independent variable.

It states results are due to chance and are not significant in supporting the idea being investigated.

The null hypothesis, positing no effect or relationship, is a foundational contrast to the research hypothesis in scientific inquiry. It establishes a baseline for statistical testing, promoting objectivity by initiating research from a neutral stance.

Many statistical methods are tailored to test the null hypothesis, determining the likelihood of observed results if no true effect exists.

This dual-hypothesis approach provides clarity, ensuring that research intentions are explicit, and fosters consistency across scientific studies, enhancing the standardization and interpretability of research outcomes.

Nondirectional Hypothesis

A non-directional hypothesis, also known as a two-tailed hypothesis, predicts that there is a difference or relationship between two variables but does not specify the direction of this relationship.

It merely indicates that a change or effect will occur without predicting which group will have higher or lower values.

For example, “There is a difference in performance between Group A and Group B” is a non-directional hypothesis.

Directional Hypothesis

A directional (one-tailed) hypothesis predicts the nature of the effect of the independent variable on the dependent variable. It predicts in which direction the change will take place. (i.e., greater, smaller, less, more)

It specifies whether one variable is greater, lesser, or different from another, rather than just indicating that there’s a difference without specifying its nature.

For example, “Exercise increases weight loss” is a directional hypothesis.

Falsifiability

The Falsification Principle, proposed by Karl Popper , is a way of demarcating science from non-science. It suggests that for a theory or hypothesis to be considered scientific, it must be testable and irrefutable.

Falsifiability emphasizes that scientific claims shouldn’t just be confirmable but should also have the potential to be proven wrong.

It means that there should exist some potential evidence or experiment that could prove the proposition false.

However many confirming instances exist for a theory, it only takes one counter observation to falsify it. For example, the hypothesis that “all swans are white,” can be falsified by observing a black swan.

For Popper, science should attempt to disprove a theory rather than attempt to continually provide evidence to support a research hypothesis.

Can a Hypothesis be Proven?

Hypotheses make probabilistic predictions. They state the expected outcome if a particular relationship exists. However, a study result supporting a hypothesis does not definitively prove it is true.

All studies have limitations. There may be unknown confounding factors or issues that limit the certainty of conclusions. Additional studies may yield different results.

In science, hypotheses can realistically only be supported with some degree of confidence, not proven. The process of science is to incrementally accumulate evidence for and against hypothesized relationships in an ongoing pursuit of better models and explanations that best fit the empirical data. But hypotheses remain open to revision and rejection if that is where the evidence leads.

• Disproving a hypothesis is definitive. Solid disconfirmatory evidence will falsify a hypothesis and require altering or discarding it based on the evidence.
• However, confirming evidence is always open to revision. Other explanations may account for the same results, and additional or contradictory evidence may emerge over time.

We can never 100% prove the alternative hypothesis. Instead, we see if we can disprove, or reject the null hypothesis.

If we reject the null hypothesis, this doesn’t mean that our alternative hypothesis is correct but does support the alternative/experimental hypothesis.

Upon analysis of the results, an alternative hypothesis can be rejected or supported, but it can never be proven to be correct. We must avoid any reference to results proving a theory as this implies 100% certainty, and there is always a chance that evidence may exist which could refute a theory.

How to Write a Hypothesis

• Identify variables . The researcher manipulates the independent variable and the dependent variable is the measured outcome.
• Operationalized the variables being investigated . Operationalization of a hypothesis refers to the process of making the variables physically measurable or testable, e.g. if you are about to study aggression, you might count the number of punches given by participants.
• Decide on a direction for your prediction . If there is evidence in the literature to support a specific effect of the independent variable on the dependent variable, write a directional (one-tailed) hypothesis. If there are limited or ambiguous findings in the literature regarding the effect of the independent variable on the dependent variable, write a non-directional (two-tailed) hypothesis.
• Make it Testable : Ensure your hypothesis can be tested through experimentation or observation. It should be possible to prove it false (principle of falsifiability).
• Clear & concise language . A strong hypothesis is concise (typically one to two sentences long), and formulated using clear and straightforward language, ensuring it’s easily understood and testable.

Consider a hypothesis many teachers might subscribe to: students work better on Monday morning than on Friday afternoon (IV=Day, DV= Standard of work).

Now, if we decide to study this by giving the same group of students a lesson on a Monday morning and a Friday afternoon and then measuring their immediate recall of the material covered in each session, we would end up with the following:

• The alternative hypothesis states that students will recall significantly more information on a Monday morning than on a Friday afternoon.
• The null hypothesis states that there will be no significant difference in the amount recalled on a Monday morning compared to a Friday afternoon. Any difference will be due to chance or confounding factors.

More Examples

• Memory : Participants exposed to classical music during study sessions will recall more items from a list than those who studied in silence.
• Social Psychology : Individuals who frequently engage in social media use will report higher levels of perceived social isolation compared to those who use it infrequently.
• Developmental Psychology : Children who engage in regular imaginative play have better problem-solving skills than those who don’t.
• Clinical Psychology : Cognitive-behavioral therapy will be more effective in reducing symptoms of anxiety over a 6-month period compared to traditional talk therapy.
• Cognitive Psychology : Individuals who multitask between various electronic devices will have shorter attention spans on focused tasks than those who single-task.
• Health Psychology : Patients who practice mindfulness meditation will experience lower levels of chronic pain compared to those who don’t meditate.
• Organizational Psychology : Employees in open-plan offices will report higher levels of stress than those in private offices.
• Behavioral Psychology : Rats rewarded with food after pressing a lever will press it more frequently than rats who receive no reward.

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Meaning of hypothesis in English

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• abstraction
• afterthought
• anthropocentrism
• anti-Darwinian
• exceptionalism
• foundation stone
• great minds think alike idiom
• non-dogmatic
• non-empirical
• non-material
• non-practical
• social Darwinism
• supersensible
• the domino theory

hypothesis | American Dictionary

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What Is A Research (Scientific) Hypothesis? A plain-language explainer + examples

By:  Derek Jansen (MBA)  | Reviewed By: Dr Eunice Rautenbach | June 2020

If you’re new to the world of research, or it’s your first time writing a dissertation or thesis, you’re probably noticing that the words “research hypothesis” and “scientific hypothesis” are used quite a bit, and you’re wondering what they mean in a research context .

“Hypothesis” is one of those words that people use loosely, thinking they understand what it means. However, it has a very specific meaning within academic research. So, it’s important to understand the exact meaning before you start hypothesizing. 

Research Hypothesis 101

• What is a hypothesis ?
• What is a research hypothesis (scientific hypothesis)?
• Requirements for a research hypothesis
• Definition of a research hypothesis
• The null hypothesis

What is a hypothesis?

Let’s start with the general definition of a hypothesis (not a research hypothesis or scientific hypothesis), according to the Cambridge Dictionary:

Hypothesis: an idea or explanation for something that is based on known facts but has not yet been proved.

In other words, it’s a statement that provides an explanation for why or how something works, based on facts (or some reasonable assumptions), but that has not yet been specifically tested . For example, a hypothesis might look something like this:

Hypothesis: sleep impacts academic performance.

This statement predicts that academic performance will be influenced by the amount and/or quality of sleep a student engages in – sounds reasonable, right? It’s based on reasonable assumptions , underpinned by what we currently know about sleep and health (from the existing literature). So, loosely speaking, we could call it a hypothesis, at least by the dictionary definition.

But that’s not good enough…

Unfortunately, that’s not quite sophisticated enough to describe a research hypothesis (also sometimes called a scientific hypothesis), and it wouldn’t be acceptable in a dissertation, thesis or research paper . In the world of academic research, a statement needs a few more criteria to constitute a true research hypothesis .

What is a research hypothesis?

A research hypothesis (also called a scientific hypothesis) is a statement about the expected outcome of a study (for example, a dissertation or thesis). To constitute a quality hypothesis, the statement needs to have three attributes – specificity , clarity and testability .

Let’s take a look at these more closely.

Need a helping hand?

Hypothesis Essential #1: Specificity & Clarity

A good research hypothesis needs to be extremely clear and articulate about both what’ s being assessed (who or what variables are involved ) and the expected outcome (for example, a difference between groups, a relationship between variables, etc.).

Let’s stick with our sleepy students example and look at how this statement could be more specific and clear.

Hypothesis: Students who sleep at least 8 hours per night will, on average, achieve higher grades in standardised tests than students who sleep less than 8 hours a night.

As you can see, the statement is very specific as it identifies the variables involved (sleep hours and test grades), the parties involved (two groups of students), as well as the predicted relationship type (a positive relationship). There’s no ambiguity or uncertainty about who or what is involved in the statement, and the expected outcome is clear.

Contrast that to the original hypothesis we looked at – “Sleep impacts academic performance” – and you can see the difference. “Sleep” and “academic performance” are both comparatively vague , and there’s no indication of what the expected relationship direction is (more sleep or less sleep). As you can see, specificity and clarity are key.

Hypothesis Essential #2: Testability (Provability)

A statement must be testable to qualify as a research hypothesis. In other words, there needs to be a way to prove (or disprove) the statement. If it’s not testable, it’s not a hypothesis – simple as that.

For example, consider the hypothesis we mentioned earlier:

Hypothesis: Students who sleep at least 8 hours per night will, on average, achieve higher grades in standardised tests than students who sleep less than 8 hours a night.  

We could test this statement by undertaking a quantitative study involving two groups of students, one that gets 8 or more hours of sleep per night for a fixed period, and one that gets less. We could then compare the standardised test results for both groups to see if there’s a statistically significant difference. 

Again, if you compare this to the original hypothesis we looked at – “Sleep impacts academic performance” – you can see that it would be quite difficult to test that statement, primarily because it isn’t specific enough. How much sleep? By who? What type of academic performance?

So, remember the mantra – if you can’t test it, it’s not a hypothesis 🙂

Defining A Research Hypothesis

You’re still with us? Great! Let’s recap and pin down a clear definition of a hypothesis.

A research hypothesis (or scientific hypothesis) is a statement about an expected relationship between variables, or explanation of an occurrence, that is clear, specific and testable.

So, when you write up hypotheses for your dissertation or thesis, make sure that they meet all these criteria. If you do, you’ll not only have rock-solid hypotheses but you’ll also ensure a clear focus for your entire research project.

What about the null hypothesis?

You may have also heard the terms null hypothesis , alternative hypothesis, or H-zero thrown around. At a simple level, the null hypothesis is the counter-proposal to the original hypothesis.

For example, if the hypothesis predicts that there is a relationship between two variables (for example, sleep and academic performance), the null hypothesis would predict that there is no relationship between those variables.

At a more technical level, the null hypothesis proposes that no statistical significance exists in a set of given observations and that any differences are due to chance alone.

And there you have it – hypotheses in a nutshell. 

If you have any questions, be sure to leave a comment below and we’ll do our best to help you. If you need hands-on help developing and testing your hypotheses, consider our private coaching service , where we hold your hand through the research journey.

Psst... there’s more!

This post was based on one of our popular Research Bootcamps . If you're working on a research project, you'll definitely want to check this out ...

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16 Comments

Very useful information. I benefit more from getting more information in this regard.

Very great insight,educative and informative. Please give meet deep critics on many research data of public international Law like human rights, environment, natural resources, law of the sea etc

In a book I read a distinction is made between null, research, and alternative hypothesis. As far as I understand, alternative and research hypotheses are the same. Can you please elaborate? Best Afshin

This is a self explanatory, easy going site. I will recommend this to my friends and colleagues.

Very good definition. How can I cite your definition in my thesis? Thank you. Is nul hypothesis compulsory in a research?

It’s a counter-proposal to be proven as a rejection

Please what is the difference between alternate hypothesis and research hypothesis?

It is a very good explanation. However, it limits hypotheses to statistically tasteable ideas. What about for qualitative researches or other researches that involve quantitative data that don’t need statistical tests?

In qualitative research, one typically uses propositions, not hypotheses.

could you please elaborate it more

I’ve benefited greatly from these notes, thank you.

This is very helpful

well articulated ideas are presented here, thank you for being reliable sources of information

Excellent. Thanks for being clear and sound about the research methodology and hypothesis (quantitative research)

I have only a simple question regarding the null hypothesis. – Is the null hypothesis (Ho) known as the reversible hypothesis of the alternative hypothesis (H1? – How to test it in academic research?

this is very important note help me much more

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Definition of hypothesis noun from the Oxford Advanced American Dictionary

• formulate/advance a theory/hypothesis
• build/construct/create/develop a simple/theoretical/mathematical model
• develop/establish/provide/use a theoretical/conceptual framework/an algorithm
• advance/argue/develop the thesis that…
• explore an idea/a concept/a hypothesis
• make a prediction/an inference
• base a prediction/your calculations on something
• investigate/evaluate/accept/challenge/reject a theory/hypothesis/model
• design an experiment/a questionnaire/a study/a test
• do research/an experiment/an analysis
• make observations/calculations
• take/record measurements
• carry out/conduct/perform an experiment/a test/a longitudinal study/observations/clinical trials
• run an experiment/a simulation/clinical trials
• repeat an experiment/a test/an analysis
• replicate a study/the results/the findings
• observe/study/examine/investigate/assess a pattern/a process/a behavior
• fund/support the research/project/study
• seek/provide/get/secure funding for research
• collect/gather/extract data/information
• yield data/evidence/similar findings/the same results
• analyze/examine the data/soil samples/a specimen
• consider/compare/interpret the results/findings
• fit the data/model
• confirm/support/verify a prediction/a hypothesis/the results/the findings
• prove a conjecture/hypothesis/theorem
• draw/make/reach the same conclusions
• read/review the records/literature
• describe/report an experiment/a study
• present/publish/summarize the results/findings
• present/publish/read/review/cite a paper in a scientific journal

Want to learn more?

Find out which words work together and produce more natural-sounding English with the Oxford Collocations Dictionary app. Try it for free as part of the Oxford Advanced Learner’s Dictionary app.

What is Hypothesis?

A hypothesis is a suggested idea or plan that has little proof, meant to lead to more study. It’s mainly a smart guess or suggested answer to a problem that can be checked through study and trial. In science work, we make guesses called hypotheses to try and figure out what will happen in tests or watching. These are not sure things but rather ideas that can be proved or disproved based on real-life proofs. A good theory is clear and can be tested and found wrong if the proof doesn’t support it.

Hypothesis Meaning

A hypothesis is a proposed statement that is testable and is given for something that happens or observed.

• It is made using what we already know and have seen, and it’s the basis for scientific research.
• A clear guess tells us what we think will happen in an experiment or study.
• It’s a testable clue that can be proven true or wrong with real-life facts and checking it out carefully.
• It usually looks like a “if-then” rule, showing the expected cause and effect relationship between what’s being studied.

Characteristics of Hypothesis

Here are some key characteristics of a hypothesis:

• Testable: An idea (hypothesis) should be made so it can be tested and proven true through doing experiments or watching. It should show a clear connection between things.
• Specific: It needs to be easy and on target, talking about a certain part or connection between things in a study.
• Falsifiable: A good guess should be able to show it’s wrong. This means there must be a chance for proof or seeing something that goes against the guess.
• Logical and Rational: It should be based on things we know now or have seen, giving a reasonable reason that fits with what we already know.
• Predictive: A guess often tells what to expect from an experiment or observation. It gives a guide for what someone might see if the guess is right.
• Concise: It should be short and clear, showing the suggested link or explanation simply without extra confusion.
• Grounded in Research: A guess is usually made from before studies, ideas or watching things. It comes from a deep understanding of what is already known in that area.
• Flexible: A guess helps in the research but it needs to change or fix when new information comes up.
• Relevant: It should be related to the question or problem being studied, helping to direct what the research is about.
• Empirical: Hypotheses come from observations and can be tested using methods based on real-world experiences.

Sources of Hypothesis

Hypotheses can come from different places based on what you’re studying and the kind of research. Here are some common sources from which hypotheses may originate:

• Existing Theories: Often, guesses come from well-known science ideas. These ideas may show connections between things or occurrences that scientists can look into more.
• Observation and Experience: Watching something happen or having personal experiences can lead to guesses. We notice odd things or repeat events in everyday life and experiments. This can make us think of guesses called hypotheses.
• Previous Research: Using old studies or discoveries can help come up with new ideas. Scientists might try to expand or question current findings, making guesses that further study old results.
• Literature Review: Looking at books and research in a subject can help make guesses. Noticing missing parts or mismatches in previous studies might make researchers think up guesses to deal with these spots.
• Problem Statement or Research Question: Often, ideas come from questions or problems in the study. Making clear what needs to be looked into can help create ideas that tackle certain parts of the issue.
• Analogies or Comparisons: Making comparisons between similar things or finding connections from related areas can lead to theories. Understanding from other fields could create new guesses in a different situation.
• Hunches and Speculation: Sometimes, scientists might get a gut feeling or make guesses that help create ideas to test. Though these may not have proof at first, they can be a beginning for looking deeper.
• Technology and Innovations: New technology or tools might make guesses by letting us look at things that were hard to study before.
• Personal Interest and Curiosity: People’s curiosity and personal interests in a topic can help create guesses. Scientists could make guesses based on their own likes or love for a subject.

Types of Hypothesis

Here are some common types of hypotheses:

Simple Hypothesis

Complex hypothesis, directional hypothesis.

• Non-directional Hypothesis

Null Hypothesis (H0)

Alternative hypothesis (h1 or ha), statistical hypothesis, research hypothesis, associative hypothesis, causal hypothesis.

Simple Hypothesis guesses a connection between two things. It says that there is a connection or difference between variables, but it doesn’t tell us which way the relationship goes.

Complex Hypothesis tells us what will happen when more than two things are connected. It looks at how different things interact and may be linked together.

Directional Hypothesis says how one thing is related to another. For example, it guesses that one thing will help or hurt another thing.

Non-Directional Hypothesis

Non-Directional Hypothesis are the one that don’t say how the relationship between things will be. They just say that there is a connection, without telling which way it goes.

Null hypothesis is a statement that says there’s no connection or difference between different things. It implies that any seen impacts are because of luck or random changes in the information.

Alternative Hypothesis is different from the null hypothesis and shows that there’s a big connection or gap between variables. Scientists want to say no to the null hypothesis and choose the alternative one.

Statistical Hypotheis are used in math testing and include making ideas about what groups or bits of them look like. You aim to get information or test certain things using these top-level, common words only.

Research Hypothesis comes from the research question and tells what link is expected between things or factors. It leads the study and chooses where to look more closely.

Associative Hypotheis guesses that there is a link or connection between things without really saying it caused them. It means that when one thing changes, it is connected to another thing changing.

Causal Hypothesis are different from other ideas because they say that one thing causes another. This means there’s a cause and effect relationship between variables involved in the situation. They say that when one thing changes, it directly makes another thing change.

Hypothesis Examples

Following are the examples of hypotheses based on their types:

Simple Hypothesis Example

• Studying more can help you do better on tests.
• Getting more sun makes people have higher amounts of vitamin D.

Complex Hypothesis Example

• How rich you are, how easy it is to get education and healthcare greatly affects the number of years people live.
• A new medicine’s success relies on the amount used, how old a person is who takes it and their genes.

Directional Hypothesis Example

• Drinking more sweet drinks is linked to a higher body weight score.
• Too much stress makes people less productive at work.

Non-directional Hypothesis Example

• Drinking caffeine can affect how well you sleep.
• People often like different kinds of music based on their gender.
• The average test scores of Group A and Group B are not much different.
• There is no connection between using a certain fertilizer and how much it helps crops grow.

Alternative Hypothesis (Ha)

• Patients on Diet A have much different cholesterol levels than those following Diet B.
• Exposure to a certain type of light can change how plants grow compared to normal sunlight.
• The average smarts score of kids in a certain school area is 100.
• The usual time it takes to finish a job using Method A is the same as with Method B.
• Having more kids go to early learning classes helps them do better in school when they get older.
• Using specific ways of talking affects how much customers get involved in marketing activities.
• Regular exercise helps to lower the chances of heart disease.
• Going to school more can help people make more money.
• Playing violent video games makes teens more likely to act aggressively.
• Less clean air directly impacts breathing health in city populations.

Functions of Hypothesis

Hypotheses have many important jobs in the process of scientific research. Here are the key functions of hypotheses:

• Guiding Research: Hypotheses give a clear and exact way for research. They act like guides, showing the predicted connections or results that scientists want to study.
• Formulating Research Questions: Research questions often create guesses. They assist in changing big questions into particular, checkable things. They guide what the study should be focused on.
• Setting Clear Objectives: Hypotheses set the goals of a study by saying what connections between variables should be found. They set the targets that scientists try to reach with their studies.
• Testing Predictions: Theories guess what will happen in experiments or observations. By doing tests in a planned way, scientists can check if what they see matches the guesses made by their ideas.
• Providing Structure: Theories give structure to the study process by arranging thoughts and ideas. They aid scientists in thinking about connections between things and plan experiments to match.
• Focusing Investigations: Hypotheses help scientists focus on certain parts of their study question by clearly saying what they expect links or results to be. This focus makes the study work better.
• Facilitating Communication: Theories help scientists talk to each other effectively. Clearly made guesses help scientists to tell others what they plan, how they will do it and the results expected. This explains things well with colleagues in a wide range of audiences.
• Generating Testable Statements: A good guess can be checked, which means it can be looked at carefully or tested by doing experiments. This feature makes sure that guesses add to the real information used in science knowledge.
• Promoting Objectivity: Guesses give a clear reason for study that helps guide the process while reducing personal bias. They motivate scientists to use facts and data as proofs or disprovals for their proposed answers.
• Driving Scientific Progress: Making, trying out and adjusting ideas is a cycle. Even if a guess is proven right or wrong, the information learned helps to grow knowledge in one specific area.

How Hypothesis help in Scientific Research?

Researchers use hypotheses to put down their thoughts directing how the experiment would take place. Following are the steps that are involved in the scientific method:

• Initiating Investigations: Hypotheses are the beginning of science research. They come from watching, knowing what’s already known or asking questions. This makes scientists make certain explanations that need to be checked with tests.
• Formulating Research Questions: Ideas usually come from bigger questions in study. They help scientists make these questions more exact and testable, guiding the study’s main point.
• Setting Clear Objectives: Hypotheses set the goals of a study by stating what we think will happen between different things. They set the goals that scientists want to reach by doing their studies.
• Designing Experiments and Studies: Assumptions help plan experiments and watchful studies. They assist scientists in knowing what factors to measure, the techniques they will use and gather data for a proposed reason.
• Testing Predictions: Ideas guess what will happen in experiments or observations. By checking these guesses carefully, scientists can see if the seen results match up with what was predicted in each hypothesis.
• Analysis and Interpretation of Data: Hypotheses give us a way to study and make sense of information. Researchers look at what they found and see if it matches the guesses made in their theories. They decide if the proof backs up or disagrees with these suggested reasons why things are happening as expected.
• Encouraging Objectivity: Hypotheses help make things fair by making sure scientists use facts and information to either agree or disagree with their suggested reasons. They lessen personal preferences by needing proof from experience.
• Iterative Process: People either agree or disagree with guesses, but they still help the ongoing process of science. Findings from testing ideas make us ask new questions, improve those ideas and do more tests. It keeps going on in the work of science to keep learning things.

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Summary – Hypothesis

A hypothesis is a testable statement serving as an initial explanation for phenomena, based on observations, theories, or existing knowledge. It acts as a guiding light for scientific research, proposing potential relationships between variables that can be empirically tested through experiments and observations. The hypothesis must be specific, testable, falsifiable, and grounded in prior research or observation, laying out a predictive, if-then scenario that details a cause-and-effect relationship. It originates from various sources including existing theories, observations, previous research, and even personal curiosity, leading to different types, such as simple, complex, directional, non-directional, null, and alternative hypotheses, each serving distinct roles in research methodology. The hypothesis not only guides the research process by shaping objectives and designing experiments but also facilitates objective analysis and interpretation of data, ultimately driving scientific progress through a cycle of testing, validation, and refinement.

FAQs on Hypothesis

What is a hypothesis.

A guess is a possible explanation or forecast that can be checked by doing research and experiments.

What are Components of a Hypothesis?

The components of a Hypothesis are Independent Variable, Dependent Variable, Relationship between Variables, Directionality etc.

What makes a Good Hypothesis?

Testability, Falsifiability, Clarity and Precision, Relevance are some parameters that makes a Good Hypothesis

Can a Hypothesis be Proven True?

You cannot prove conclusively that most hypotheses are true because it’s generally impossible to examine all possible cases for exceptions that would disprove them.

How are Hypotheses Tested?

Hypothesis testing is used to assess the plausibility of a hypothesis by using sample data

Can Hypotheses change during Research?

Yes, you can change or improve your ideas based on new information discovered during the research process.

What is the Role of a Hypothesis in Scientific Research?

Hypotheses are used to support scientific research and bring about advancements in knowledge.

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Hypothesis Testing

Hypothesis testing is a tool for making statistical inferences about the population data. It is an analysis tool that tests assumptions and determines how likely something is within a given standard of accuracy. Hypothesis testing provides a way to verify whether the results of an experiment are valid.

A null hypothesis and an alternative hypothesis are set up before performing the hypothesis testing. This helps to arrive at a conclusion regarding the sample obtained from the population. In this article, we will learn more about hypothesis testing, its types, steps to perform the testing, and associated examples.

What is Hypothesis Testing in Statistics?

Hypothesis testing uses sample data from the population to draw useful conclusions regarding the population probability distribution . It tests an assumption made about the data using different types of hypothesis testing methodologies. The hypothesis testing results in either rejecting or not rejecting the null hypothesis.

Hypothesis Testing Definition

Hypothesis testing can be defined as a statistical tool that is used to identify if the results of an experiment are meaningful or not. It involves setting up a null hypothesis and an alternative hypothesis. These two hypotheses will always be mutually exclusive. This means that if the null hypothesis is true then the alternative hypothesis is false and vice versa. An example of hypothesis testing is setting up a test to check if a new medicine works on a disease in a more efficient manner.

Null Hypothesis

The null hypothesis is a concise mathematical statement that is used to indicate that there is no difference between two possibilities. In other words, there is no difference between certain characteristics of data. This hypothesis assumes that the outcomes of an experiment are based on chance alone. It is denoted as \(H_{0}\). Hypothesis testing is used to conclude if the null hypothesis can be rejected or not. Suppose an experiment is conducted to check if girls are shorter than boys at the age of 5. The null hypothesis will say that they are the same height.

Alternative Hypothesis

The alternative hypothesis is an alternative to the null hypothesis. It is used to show that the observations of an experiment are due to some real effect. It indicates that there is a statistical significance between two possible outcomes and can be denoted as \(H_{1}\) or \(H_{a}\). For the above-mentioned example, the alternative hypothesis would be that girls are shorter than boys at the age of 5.

Hypothesis Testing P Value

In hypothesis testing, the p value is used to indicate whether the results obtained after conducting a test are statistically significant or not. It also indicates the probability of making an error in rejecting or not rejecting the null hypothesis.This value is always a number between 0 and 1. The p value is compared to an alpha level, \(\alpha\) or significance level. The alpha level can be defined as the acceptable risk of incorrectly rejecting the null hypothesis. The alpha level is usually chosen between 1% to 5%.

Hypothesis Testing Critical region

All sets of values that lead to rejecting the null hypothesis lie in the critical region. Furthermore, the value that separates the critical region from the non-critical region is known as the critical value.

Hypothesis Testing Formula

Depending upon the type of data available and the size, different types of hypothesis testing are used to determine whether the null hypothesis can be rejected or not. The hypothesis testing formula for some important test statistics are given below:

• z = \(\frac{\overline{x}-\mu}{\frac{\sigma}{\sqrt{n}}}\). \(\overline{x}\) is the sample mean, \(\mu\) is the population mean, \(\sigma\) is the population standard deviation and n is the size of the sample.
• t = \(\frac{\overline{x}-\mu}{\frac{s}{\sqrt{n}}}\). s is the sample standard deviation.
• \(\chi ^{2} = \sum \frac{(O_{i}-E_{i})^{2}}{E_{i}}\). \(O_{i}\) is the observed value and \(E_{i}\) is the expected value.

We will learn more about these test statistics in the upcoming section.

Types of Hypothesis Testing

Selecting the correct test for performing hypothesis testing can be confusing. These tests are used to determine a test statistic on the basis of which the null hypothesis can either be rejected or not rejected. Some of the important tests used for hypothesis testing are given below.

Hypothesis Testing Z Test

A z test is a way of hypothesis testing that is used for a large sample size (n ≥ 30). It is used to determine whether there is a difference between the population mean and the sample mean when the population standard deviation is known. It can also be used to compare the mean of two samples. It is used to compute the z test statistic. The formulas are given as follows:

• One sample: z = \(\frac{\overline{x}-\mu}{\frac{\sigma}{\sqrt{n}}}\).
• Two samples: z = \(\frac{(\overline{x_{1}}-\overline{x_{2}})-(\mu_{1}-\mu_{2})}{\sqrt{\frac{\sigma_{1}^{2}}{n_{1}}+\frac{\sigma_{2}^{2}}{n_{2}}}}\).

Hypothesis Testing t Test

The t test is another method of hypothesis testing that is used for a small sample size (n < 30). It is also used to compare the sample mean and population mean. However, the population standard deviation is not known. Instead, the sample standard deviation is known. The mean of two samples can also be compared using the t test.

• One sample: t = \(\frac{\overline{x}-\mu}{\frac{s}{\sqrt{n}}}\).
• Two samples: t = \(\frac{(\overline{x_{1}}-\overline{x_{2}})-(\mu_{1}-\mu_{2})}{\sqrt{\frac{s_{1}^{2}}{n_{1}}+\frac{s_{2}^{2}}{n_{2}}}}\).

Hypothesis Testing Chi Square

The Chi square test is a hypothesis testing method that is used to check whether the variables in a population are independent or not. It is used when the test statistic is chi-squared distributed.

One Tailed Hypothesis Testing

One tailed hypothesis testing is done when the rejection region is only in one direction. It can also be known as directional hypothesis testing because the effects can be tested in one direction only. This type of testing is further classified into the right tailed test and left tailed test.

Right Tailed Hypothesis Testing

The right tail test is also known as the upper tail test. This test is used to check whether the population parameter is greater than some value. The null and alternative hypotheses for this test are given as follows:

\(H_{0}\): The population parameter is ≤ some value

\(H_{1}\): The population parameter is > some value.

If the test statistic has a greater value than the critical value then the null hypothesis is rejected

Left Tailed Hypothesis Testing

The left tail test is also known as the lower tail test. It is used to check whether the population parameter is less than some value. The hypotheses for this hypothesis testing can be written as follows:

\(H_{0}\): The population parameter is ≥ some value

\(H_{1}\): The population parameter is < some value.

The null hypothesis is rejected if the test statistic has a value lesser than the critical value.

Two Tailed Hypothesis Testing

In this hypothesis testing method, the critical region lies on both sides of the sampling distribution. It is also known as a non - directional hypothesis testing method. The two-tailed test is used when it needs to be determined if the population parameter is assumed to be different than some value. The hypotheses can be set up as follows:

\(H_{0}\): the population parameter = some value

\(H_{1}\): the population parameter ≠ some value

The null hypothesis is rejected if the test statistic has a value that is not equal to the critical value.

Hypothesis Testing Steps

Hypothesis testing can be easily performed in five simple steps. The most important step is to correctly set up the hypotheses and identify the right method for hypothesis testing. The basic steps to perform hypothesis testing are as follows:

• Step 1: Set up the null hypothesis by correctly identifying whether it is the left-tailed, right-tailed, or two-tailed hypothesis testing.
• Step 2: Set up the alternative hypothesis.
• Step 3: Choose the correct significance level, \(\alpha\), and find the critical value.
• Step 4: Calculate the correct test statistic (z, t or \(\chi\)) and p-value.
• Step 5: Compare the test statistic with the critical value or compare the p-value with \(\alpha\) to arrive at a conclusion. In other words, decide if the null hypothesis is to be rejected or not.

Hypothesis Testing Example

The best way to solve a problem on hypothesis testing is by applying the 5 steps mentioned in the previous section. Suppose a researcher claims that the mean average weight of men is greater than 100kgs with a standard deviation of 15kgs. 30 men are chosen with an average weight of 112.5 Kgs. Using hypothesis testing, check if there is enough evidence to support the researcher's claim. The confidence interval is given as 95%.

Step 1: This is an example of a right-tailed test. Set up the null hypothesis as \(H_{0}\): \(\mu\) = 100.

Step 2: The alternative hypothesis is given by \(H_{1}\): \(\mu\) > 100.

Step 3: As this is a one-tailed test, \(\alpha\) = 100% - 95% = 5%. This can be used to determine the critical value.

1 - \(\alpha\) = 1 - 0.05 = 0.95

0.95 gives the required area under the curve. Now using a normal distribution table, the area 0.95 is at z = 1.645. A similar process can be followed for a t-test. The only additional requirement is to calculate the degrees of freedom given by n - 1.

Step 4: Calculate the z test statistic. This is because the sample size is 30. Furthermore, the sample and population means are known along with the standard deviation.

z = \(\frac{\overline{x}-\mu}{\frac{\sigma}{\sqrt{n}}}\).

\(\mu\) = 100, \(\overline{x}\) = 112.5, n = 30, \(\sigma\) = 15

z = \(\frac{112.5-100}{\frac{15}{\sqrt{30}}}\) = 4.56

Step 5: Conclusion. As 4.56 > 1.645 thus, the null hypothesis can be rejected.

Hypothesis Testing and Confidence Intervals

Confidence intervals form an important part of hypothesis testing. This is because the alpha level can be determined from a given confidence interval. Suppose a confidence interval is given as 95%. Subtract the confidence interval from 100%. This gives 100 - 95 = 5% or 0.05. This is the alpha value of a one-tailed hypothesis testing. To obtain the alpha value for a two-tailed hypothesis testing, divide this value by 2. This gives 0.05 / 2 = 0.025.

Related Articles:

• Probability and Statistics
• Data Handling

Important Notes on Hypothesis Testing

• Hypothesis testing is a technique that is used to verify whether the results of an experiment are statistically significant.
• It involves the setting up of a null hypothesis and an alternate hypothesis.
• There are three types of tests that can be conducted under hypothesis testing - z test, t test, and chi square test.
• Hypothesis testing can be classified as right tail, left tail, and two tail tests.

Examples on Hypothesis Testing

• Example 1: The average weight of a dumbbell in a gym is 90lbs. However, a physical trainer believes that the average weight might be higher. A random sample of 5 dumbbells with an average weight of 110lbs and a standard deviation of 18lbs. Using hypothesis testing check if the physical trainer's claim can be supported for a 95% confidence level. Solution: As the sample size is lesser than 30, the t-test is used. \(H_{0}\): \(\mu\) = 90, \(H_{1}\): \(\mu\) > 90 \(\overline{x}\) = 110, \(\mu\) = 90, n = 5, s = 18. \(\alpha\) = 0.05 Using the t-distribution table, the critical value is 2.132 t = \(\frac{\overline{x}-\mu}{\frac{s}{\sqrt{n}}}\) t = 2.484 As 2.484 > 2.132, the null hypothesis is rejected. Answer: The average weight of the dumbbells may be greater than 90lbs
• Example 2: The average score on a test is 80 with a standard deviation of 10. With a new teaching curriculum introduced it is believed that this score will change. On random testing, the score of 38 students, the mean was found to be 88. With a 0.05 significance level, is there any evidence to support this claim? Solution: This is an example of two-tail hypothesis testing. The z test will be used. \(H_{0}\): \(\mu\) = 80, \(H_{1}\): \(\mu\) ≠ 80 \(\overline{x}\) = 88, \(\mu\) = 80, n = 36, \(\sigma\) = 10. \(\alpha\) = 0.05 / 2 = 0.025 The critical value using the normal distribution table is 1.96 z = \(\frac{\overline{x}-\mu}{\frac{\sigma}{\sqrt{n}}}\) z = \(\frac{88-80}{\frac{10}{\sqrt{36}}}\) = 4.8 As 4.8 > 1.96, the null hypothesis is rejected. Answer: There is a difference in the scores after the new curriculum was introduced.
• Example 3: The average score of a class is 90. However, a teacher believes that the average score might be lower. The scores of 6 students were randomly measured. The mean was 82 with a standard deviation of 18. With a 0.05 significance level use hypothesis testing to check if this claim is true. Solution: The t test will be used. \(H_{0}\): \(\mu\) = 90, \(H_{1}\): \(\mu\) < 90 \(\overline{x}\) = 110, \(\mu\) = 90, n = 6, s = 18 The critical value from the t table is -2.015 t = \(\frac{\overline{x}-\mu}{\frac{s}{\sqrt{n}}}\) t = \(\frac{82-90}{\frac{18}{\sqrt{6}}}\) t = -1.088 As -1.088 > -2.015, we fail to reject the null hypothesis. Answer: There is not enough evidence to support the claim.

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FAQs on Hypothesis Testing

What is hypothesis testing.

Hypothesis testing in statistics is a tool that is used to make inferences about the population data. It is also used to check if the results of an experiment are valid.

What is the z Test in Hypothesis Testing?

The z test in hypothesis testing is used to find the z test statistic for normally distributed data . The z test is used when the standard deviation of the population is known and the sample size is greater than or equal to 30.

What is the t Test in Hypothesis Testing?

The t test in hypothesis testing is used when the data follows a student t distribution . It is used when the sample size is less than 30 and standard deviation of the population is not known.

What is the formula for z test in Hypothesis Testing?

The formula for a one sample z test in hypothesis testing is z = \(\frac{\overline{x}-\mu}{\frac{\sigma}{\sqrt{n}}}\) and for two samples is z = \(\frac{(\overline{x_{1}}-\overline{x_{2}})-(\mu_{1}-\mu_{2})}{\sqrt{\frac{\sigma_{1}^{2}}{n_{1}}+\frac{\sigma_{2}^{2}}{n_{2}}}}\).

What is the p Value in Hypothesis Testing?

The p value helps to determine if the test results are statistically significant or not. In hypothesis testing, the null hypothesis can either be rejected or not rejected based on the comparison between the p value and the alpha level.

What is One Tail Hypothesis Testing?

When the rejection region is only on one side of the distribution curve then it is known as one tail hypothesis testing. The right tail test and the left tail test are two types of directional hypothesis testing.

What is the Alpha Level in Two Tail Hypothesis Testing?

To get the alpha level in a two tail hypothesis testing divide \(\alpha\) by 2. This is done as there are two rejection regions in the curve.

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[ hahy- poth - uh -sis , hi- ]

• a proposition, or set of propositions, set forth as an explanation for the occurrence of some specified group of phenomena, either asserted merely as a provisional conjecture to guide investigation working hypothesis or accepted as highly probable in the light of established facts.
• a proposition assumed as a premise in an argument.
• the antecedent of a conditional proposition.
• a mere assumption or guess.

/ haɪˈpɒθɪsɪs /

• a suggested explanation for a group of facts or phenomena, either accepted as a basis for further verification ( working hypothesis ) or accepted as likely to be true Compare theory
• an assumption used in an argument without its being endorsed; a supposition
• an unproved theory; a conjecture

/ hī-pŏth ′ ĭ-sĭs /

, Plural hypotheses hī-pŏth ′ ĭ-sēz′

• A statement that explains or makes generalizations about a set of facts or principles, usually forming a basis for possible experiments to confirm its viability.
• plur. hypotheses (heye- poth -uh-seez) In science, a statement of a possible explanation for some natural phenomenon. A hypothesis is tested by drawing conclusions from it; if observation and experimentation show a conclusion to be false, the hypothesis must be false. ( See scientific method and theory .)

Discover More

Derived forms.

• hyˈpothesist , noun

Other Words From

• hy·pothe·sist noun
• counter·hy·pothe·sis noun plural counterhypotheses
• subhy·pothe·sis noun plural subhypotheses

Word History and Origins

Origin of hypothesis 1

Synonym Study

Example sentences.

Though researchers have struggled to understand exactly what contributes to this gender difference, Dr. Rohan has one hypothesis.

The leading hypothesis for the ultimate source of the Ebola virus, and where it retreats in between outbreaks, lies in bats.

In 1996, John Paul II called the Big Bang theory “more than a hypothesis.”

To be clear: There have been no double-blind or controlled studies that conclusively confirm this hair-loss hypothesis.

The bacteria-driven-ritual hypothesis ignores the huge diversity of reasons that could push someone to perform a religious ritual.

And remember it is by our hypothesis the best possible form and arrangement of that lesson.

Taken in connection with what we know of the nebulæ, the proof of Laplace's nebular hypothesis may fairly be regarded as complete.

What has become of the letter from M. de St. Mars, said to have been discovered some years ago, confirming this last hypothesis?

To admit that there had really been any communication between the dead man and the living one is also an hypothesis.

"I consider it highly probable," asserted Aunt Maria, forgetting her Scandinavian hypothesis.

Related Words

• explanation
• interpretation
• proposition
• supposition

• Math Article

Hypothesis Definition

In Statistics, the determination of the variation between the group of data due to true variation is done by hypothesis testing. The sample data are taken from the population parameter based on the assumptions. The hypothesis can be classified into various types. In this article, let us discuss the hypothesis definition, various types of hypothesis and the significance of hypothesis testing, which are explained in detail.

Hypothesis Definition in Statistics

In Statistics, a hypothesis is defined as a formal statement, which gives the explanation about the relationship between the two or more variables of the specified population. It helps the researcher to translate the given problem to a clear explanation for the outcome of the study. It clearly explains and predicts the expected outcome. It indicates the types of experimental design and directs the study of the research process.

Types of Hypothesis

The hypothesis can be broadly classified into different types. They are:

Simple Hypothesis

A simple hypothesis is a hypothesis that there exists a relationship between two variables. One is called a dependent variable, and the other is called an independent variable.

Complex Hypothesis

A complex hypothesis is used when there is a relationship between the existing variables. In this hypothesis, the dependent and independent variables are more than two.

Null Hypothesis

In the null hypothesis, there is no significant difference between the populations specified in the experiments, due to any experimental or sampling error. The null hypothesis is denoted by H 0 .

Alternative Hypothesis

In an alternative hypothesis, the simple observations are easily influenced by some random cause. It is denoted by the H a or H 1 .

Empirical Hypothesis

An empirical hypothesis is formed by the experiments and based on the evidence.

Statistical Hypothesis

In a statistical hypothesis, the statement should be logical or illogical, and the hypothesis is verified statistically.

Apart from these types of hypothesis, some other hypotheses are directional and non-directional hypothesis, associated hypothesis, casual hypothesis.

Characteristics of Hypothesis

The important characteristics of the hypothesis are:

• The hypothesis should be short and precise
• It should be specific
• A hypothesis must be related to the existing body of knowledge
• It should be capable of verification

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• Open access
• Published: 13 May 2024

SCIPAC: quantitative estimation of cell-phenotype associations

• Dailin Gan 1 ,
• Yini Zhu 2 ,
• Xin Lu 2 , 3 &
• Jun Li   ORCID: orcid.org/0000-0003-4353-5761 1  

Genome Biology volume  25 , Article number:  119 ( 2024 ) Cite this article

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Numerous algorithms have been proposed to identify cell types in single-cell RNA sequencing data, yet a fundamental problem remains: determining associations between cells and phenotypes such as cancer. We develop SCIPAC, the first algorithm that quantitatively estimates the association between each cell in single-cell data and a phenotype. SCIPAC also provides a p -value for each association and applies to data with virtually any type of phenotype. We demonstrate SCIPAC’s accuracy in simulated data. On four real cancerous or noncancerous datasets, insights from SCIPAC help interpret the data and generate new hypotheses. SCIPAC requires minimum tuning and is computationally very fast.

Single-cell RNA sequencing (scRNA-seq) technologies are revolutionizing biomedical research by providing comprehensive characterizations of diverse cell populations in heterogeneous tissues [ 1 , 2 ]. Unlike bulk RNA sequencing (RNA-seq), which measures the average expression profile of the whole tissue, scRNA-seq gives the expression profiles of thousands of individual cells in the tissue [ 3 , 4 , 5 , 6 , 7 ]. Based on this rich data, cell types may be discovered/determined in an unsupervised (e.g., [ 8 , 9 ]), semi-supervised (e.g., [ 10 , 11 , 12 , 13 ]), or supervised manner (e.g., [ 14 , 15 , 16 ]). Despite the fast development, there are still limitations with scRNA-seq technologies. Notably, the cost for each scRNA-seq experiment is still high; as a result, most scRNA-seq data are from a single or a few biological samples/tissues. Very few datasets consist of large numbers of samples with different phenotypes, e.g., cancer vs. normal. This places great difficulties in determining how a cell type contributes to a phenotype based on single-cell studies (especially if the cell type is discovered in a completely unsupervised manner or if people have limited knowledge of this cell type). For example, without having single-cell data from multiple cancer patients and multiple normal controls, it could be hard to computationally infer whether a cell type may promote or inhibit cancer development. However, such association can be critical for cancer research [ 17 ], disease diagnosis [ 18 ], cell-type targeted therapy development [ 19 ], etc.

Fortunately, this difficulty may be overcome by borrowing information from bulk RNA-seq data. Over the past decade, a considerable amount of bulk RNA-seq data from a large number of samples with different phenotypes have been accumulated and made available through databases like The Cancer Genome Atlas (TCGA) [ 20 ] and cBioPortal [ 21 , 22 ]. Data in these databases often contain comprehensive patient phenotype information, such as cancer status, cancer stages, survival status and time, and tumor metastasis. Combining single-cell data from a single or a few individuals and bulk data from a relatively large number of individuals regarding a particular phenotype can be a cost-effective way to determine how a cell type contributes to the phenotype. A recent method Scissor [ 23 ] took an essential step in this direction. It uses single-cell and bulk RNA-seq data with phenotype information to classify the cells into three discrete categories: Scissor+, Scissor−, and null cells, corresponding to cells that are positively associated, negatively associated, and not associated with the phenotype.

Here, we present a method that takes another big step in this direction, which is called Single-Cell and bulk data-based Identifier for Phenotype Associated Cells or SCIPAC for short. SCIPAC enables quantitative estimation of the strength of association between each cell in a scRNA-seq data and a phenotype, with the help of bulk RNA-seq data with phenotype information. Moreover, SCIPAC also enables the estimation of the statistical significance of the association. That is, it gives a p -value for the association between each cell and the phenotype. Furthermore, SCIPAC enables the estimation of association between cells and an ordinal phenotype (e.g., different stages of cancer), which could be informative as people may not only be interested in the emergence/existence of cancer (cancer vs. healthy, a binary problem) but also in the progression of cancer (different stages of cancer, an ordinal problem).

To study the performance of SCIPAC, we first apply SCIPAC to simulated data under three schemes. SCIPAC shows high accuracy with low false positive rates. We further show the broad applicability of SCIPAC on real datasets across various diseases, including prostate cancer, breast cancer, lung cancer, and muscular dystrophy. The association inferred by SCIPAC is highly informative. In real datasets, some cell types have definite and well-studied functions, while others are less well-understood: their functions may be disease-dependent or tissue-dependent, and they may contain different sub-types with distinct functions. In the former case, SCIPAC’s results agree with current biological knowledge. In the latter case, SCIPAC’s discoveries inspire the generation of new hypotheses regarding the roles and functions of cells under different conditions.

An overview of the SCIPAC algorithm

SCIPAC is a computational method that identifies cells in single-cell data that are associated with a given phenotype. This phenotype can be binary (e.g., cancer vs. normal), ordinal (e.g., cancer stage), continuous (e.g., quantitative traits), or survival (i.e., survival time and status). SCIPAC uses input data consisting of three parts: single-cell RNA-seq data that measures the expression of p genes in m cells, bulk RNA-seq data that measures the expression of the same set of p genes in n samples/tissues, and the statuses/values of the phenotype of the n bulk samples/tissues. The output of SCIPAC is the strength and the p -value of the association between each cell and the phenotype.

SCIPAC proposes the following definition of “association” between a cell and a phenotype: A group of cells that are likely to play a similar role in the phenotype (such as cells of a specific cell type or sub-type, cells in a particular state, cells in a cluster, cells with similar expression profiles, or cells with similar functions) is considered to be positively/negatively associated with a phenotype if an increase in their proportion within the tissue likely indicates an increased/decreased probability of the phenotype’s presence. SCIPAC assigns the same association to all cells within such a group. Taking cancer as the phenotype as an example, if increasing the proportion of a cell type indicates a higher chance of having cancer (binary), having a higher cancer stage (ordinal), or a higher hazard rate (survival), all cells in this cell type is positively associated with cancer.

The algorithm of SCIPAC follows the following four steps. First, the cells in the single-cell data are grouped into clusters according to their expression profiles. The Louvain algorithm from the Seurat package [ 24 , 25 ] is used as the default clustering algorithm, but the user may choose any clustering algorithm they prefer. Or if information of the cell types or other groupings of cells is available a priori, it may be supplied to SCIPAC as the cell clusters, and this clustering step can be skipped. In the second step, a regression model is learned from bulk gene expression data with the phenotype. Depending on the type of the phenotype, this model can be logistic regression, ordinary linear regression, proportional odds model, or Cox proportional hazards model. To achieve a higher prediction power with less variance, by default, the elastic net (a blender of Lasso and ridge regression [ 26 ]) is used to fit the model. In the third step, SCIPAC computes the association strength \(\Lambda\) between each cell cluster and the phenotype based on a mathematical formula that we derive. Finally, the p -values are computed. The association strength and its p -value between a cell cluster and the phenotype are given to all cells in the cluster.

SCIPAC requires minimum tuning. When the cell-type information is given in step 1, SCIPAC does not have any (hyper)parameter. Otherwise, the Louvain algorithm used in step 1 has a “resolution” parameter that controls the number of cell clusters: a larger resolution results in more clusters. SCIPAC inherits this parameter as its only parameter. Since SCIPAC gives the same association strength and p -value to cells from the same cluster, this parameter also determines the resolution of results provided by SCIPAC. Thus, we still call it “resolution” in SCIPAC. Because of its meaning, we recommend setting it so that the number of cell clusters given by the clustering algorithm is comparable to, or reasonably larger than, the number of cell types (or sub-types) in the data. We will see that the performance of SCIPAC is insensitive to this resolution parameter, and the default value 2.0 typically works well.

The details of the SCIPAC algorithm are given in the “ Methods ” section.

Performance in simulated data

We assess the performance of SCIPAC in simulated data under three different schemes. The first scheme is simple and consists of only three cell types. The second scheme is more complicated and consists of seven cell types, which better imitates actual scRNA-seq data. In the third scheme, we simulate cells under different cell development stages to test the performance of SCIPAC under an ordinal phenotype. Details of the simulation are given in Additional file 1.

Simulation scheme I

Under this scheme, the single-cell data consists of three cell types: one is positively associated with the phenotype, one is negatively associated, and the third is not associated (we call it “null”). Figure 1 a gives the UMAP [ 27 ] plot of the three cell types, and Fig. 1 b gives the true associations of these three cell types with the phenotype, with red, blue, and light gray denoting positive, negative, and null associations.

UMAP visualization and numeric measures of the simulated data under scheme I. All the plots in a–e  are scatterplots of the two dimensional single-cell data given by UMAP. The x and y axes represent the two dimensions, and their scales are not shown as their specific values are not directly relevant. Points in the plots represents single cells, and they are colored differently in each subplot to reflect different information/results. a  Cell types. b  True associations. The association between cell types 1, 2, and 3 and the phenotype are positive, negative, and null, respectively. c  Association strengths \(\Lambda\) given by SCIPAC under different resolutions. Red/blue represents the sign of \(\Lambda\) , and the shade gives the absolute value of \(\Lambda\) . Every cell is colored red or blue since no \(\Lambda\) is exactly zero. Below each subplot, Res stands for resolution, and K stands for the number of cell clusters given by this resolution. d   p -values given by SCIPAC. Only cells with p -value \(< 0.05\) are colored red (positive association) or blue (negative association); others are colored white. e  Results given by Scissor under different \(\alpha\) values. Red, blue, and light gray stands for Scissor+, Scissor−, and background (i.e., null) cells. f  F1 scores and g  FSC for SCIPAC and Scissor under different parameter values. For SCIPAC, each bar is the value under a resolution/number of clusters. For Scissor, each bar is the value under an \(\alpha\)

We apply SCIPAC to the simulated data. For the resolution parameter (see the “ Methods ” section), values 0.5, 1.0, and 1.5 give 3, 4, and 4 clusters, respectively, close to the actual number of cell types. They are good choices based on the guidance for choosing this parameter. To show how SCIPAC behaves under parameter misspecification, we also set the resolution up to 4.0, which gives a whopping 61 clusters. Figure 1 c and d give the association strengths \(\Lambda\) and the p -values given by four different resolutions (results under other resolutions are provided in Additional file 1: Fig. S1 and S2). In Fig. 1 c, red and blue denote positive and negative associations, respectively, and the shade of the color represents the strength of the association, i.e., the absolute value of \(\Lambda\) . Every cell is colored blue or red, as none of \(\Lambda\) is exactly zero. In Fig. 1 d, red and blue denote positive and negative associations that are statistically significant ( p -value \(< 0.05\) ). Cells whose associations are not statistically significant ( p -value \(\ge 0.05\) ) are shown in white. To avoid confusion, it is worth repeating that cells that are colored in red/blue in Fig. 1 c are shown in red/blue in Fig. 1 d only if they are statistically significant; otherwise, they are colored white in Fig. 1 d.

From Fig. 1 c, d (as well as Additional file 1: Fig. S1 and S2), it is clear that the results of SCIPAC are highly consistent under different resolution values, including both the estimated association strengths and the p -values. It is also clear that SCIPAC is highly accurate: most truly associated cells are identified as significant, and most, if not all, truly null cells are identified as null.

As the first algorithm that quantitatively estimates the association strength and the first algorithm that gives the p -value of the association, SCIPAC does not have a real competitor. A previous algorithm, Scissor, is able to classify cells into three discrete categories according to their associations with the phenotype. So, we compare SCIPAC with Scissor in respect of the ability to differentiate positively associated, negatively associated, and null cells.

Running Scissor requires tuning a parameter called \(\alpha\) , which is a number between 0 and 1 that balances the amount of regularization for the smoothness and for the sparsity of the associations. The Scissor R package does not provide a default value for this \(\alpha\) or a function to help select this value. The Scissor paper suggests the following criterion: “the number of Scissor-selected cells should not exceed a certain percentage of total cells (default 20%) in the single-cell data. In each experiment, a search on the above searching list is performed from the smallest to the largest until a value of \(\alpha\) meets the above criteria.” In practice, we have found that this criterion does not often work properly, as the truly associated cells may not compose 20% of all cells in actual data. Therefore, instead of setting \(\alpha\) to any particular value, we set \(\alpha\) values that span the whole range of \(\alpha\) to see the best possible performance of Scissor.

The performance of Scissor in our simulation data under four different \(\alpha\) values are shown in Fig. 1 e, and results under more \(\alpha\) values are shown in Additional file 1: Fig. S3. In the figures, red, blue, and light gray denote Scissor+, Scissor−, and null (called “background” in Scissor) cells, respectively. The results of Scissor have several characteristics different from SCIPAC. First, Scissor does not give the strength or statistical significance of the association, and thus the colors of the cells in the figures do not have different shades. Second, different \(\alpha\) values give very different results. Greater \(\alpha\) values generally give fewer Scissor+ and Scissor− cells, but there are additional complexities. One complexity is that the Scissor+ (or Scissor−) cells under a greater \(\alpha\) value are not a strict subset of Scissor+ (or Scissor−) cells under a smaller \(\alpha\) value. For example, the number of truly negatively associated cells detected as Scissor− increases when \(\alpha\) increases from 0.01 to 0.30. Another complexity is that the direction of the association may flip as \(\alpha\) increases. For example, most cells of cell type 2 are identified as Scissor+ under \(\alpha =0.01\) , but many of them are identified as Scissor− under larger \(\alpha\) values. Third, Scissor does not achieve high power and low false-positive rate at the same time under any \(\alpha\) . No matter what the \(\alpha\) value is, there is only a small proportion of cells from cell type 2 that are correctly identified as negatively associated, and there is always a non-negligible proportion of null cells (i.e., cells from cell type 3) that are incorrectly identified as positively or negatively associated. Fourth, Scissor+ and Scissor− cells can be close to each other in the figure, even under a large \(\alpha\) value. This means that cells with nearly identical expression profiles are detected to be associated with the phenotype in opposite directions, which can place difficulties in interpreting the results.

SCIPAC overcomes the difficulties of Scissor and gives results that are more informative (quantitative strengths with p -values), more accurate (both high power and low false-positive rate), less sensitive to the tuning parameter, and easier to interpret (cells with similar expression typically have similar associations to the phenotype).

SCIPAC’s higher accuracy in differentiating positively associated, negatively associated, and null cells than Scissors can also be measured numerically using the F1 score and the fraction of sign correctness (FSC). F1, which is the harmonic mean of precision and recall, is a commonly used measure of calling accuracy. Note that precision and recall are only defined for two-class problems, which try to classify desired signals/discoveries (so-called “positives”) against noises/trivial results (so-called “negatives”). Our case, on the other hand, is a three-class problem: positive association, negative association, and null. To compute F1, we combine the positive and negative associations and treat them as “positives,” and treat null as “negatives.” This F1 score ignores the direction of the association; thus, it alone is not enough to describe the performance of an association-detection algorithm. For example, an algorithm may have a perfect F1 score even if it incorrectly calls all negative associations positive. To measure an algorithm’s ability to determine the direction of the association, we propose a statistic called FSC, defined as the fraction of true discoveries that also have the correct direction of the association. The F1 score and FSC are numbers between 0 and 1, and higher values are preferred. A mathematical definition of these two measures is given in Additional file 1.

Figure 1 f, g show the F1 score and FSC of SCIPAC and Scissor under different values of tuning parameters. The F1 score of Scissor is between 0.2 and 0.7 under different \(\alpha\) ’s. The FSC of Scissor increases from around 0.5 to nearly 1 as \(\alpha\) increases, but Scissor does not achieve high F1 and FSC scores at the same time under any \(\alpha\) . On the other hand, the F1 score of SCIPAC is close to perfection when the resolution parameter is properly set, and it is still above 0.90 even if the resolution parameter is set too large. The FSC of SCIPAC is always above 0.96 under different resolutions. That is, SCIPAC achieves high F1 and FSC scores simultaneously under a wide range of resolutions, representing a much higher accuracy than Scissor.

Simulation scheme II

This more complicated simulation scheme has seven cell types, which are shown in Fig. 2 a. As shown in Fig. 2 b, cell types 1 and 3 are negatively associated (colored blue), 2 and 4 are positively associated (colored red), and 5, 6, and 7 are not associated (colored light gray).

UMAP visualization of the simulated data under a–g  scheme II and h–k  scheme III. a  Cell types. b  True associations. c , d  Association strengths \(\Lambda\) and p -values given by SCIPAC under the default resolution. e  Results given by Scissor under different \(\alpha\) values. f  F1 scores and g  FSC for SCIPAC and Scissor under different parameter values. h  Cell differentiation paths. The four paths have the same starting location, which is in the center, but different ending locations. This can be considered as a progenitor cell type differentiating into four specialized cell types. i  Cell differentiation steps. These steps are used to create four stages, each containing 500 steps. Thus, this plot of differentiation steps can also be viewed as the plot of true association strengths. j , k  Association strengths \(\Lambda\) and p -values given by SCIPAC under the default resolution

The association strengths and p -values given by SCIPAC under the default resolution are illustrated in Fig. 2 c, d, respectively. Results under several other resolutions are given in Additional file 1: Fig. S4 and S5. Again, we find that SCIPAC gives highly consistent results under different resolutions. SCIPAC successfully identifies three out of the four truly associated cell types. For the other truly associated cell type, cell type 1, SCIPAC correctly recognizes its association with the phenotype as negative, although the p -values are not significant enough. The F1 score is 0.85, and the FSC is greater than 0.99, as shown in Fig. 2 f, g.

The results of Scissor under four different \(\alpha\) values are given in Fig. 2 e. (More shown in Additional file 1: Fig. S6.) Under this highly challenging simulation scheme, Scissor can only identify one out of four truly associated cell types. Its F1 score is below 0.4.

Simulation scheme III

This simulation scheme is to assess the performance of SCIPAC for ordinal phenotypes. We simulate cells along four cell-differentiation paths with the same starting location but different ending locations, as shown in Fig. 2 h. These cells can be considered as a progenitor cell population differentiating into four specialized cell types. In Fig. 2 i, the “step” reflects their position in the differentiation path, with step 0 meaning the start and step 2000 meaning the end of the differentiation. Then, the “stage” is generated according to the step: cells in steps 0 \(\sim\) 500, 501 \(\sim\) 1000, 1001 \(\sim\) 1500, and 1501 \(\sim\) 2000 are assigned to stages I, II, III, and IV, respectively. This stage is treated as the ordinal phenotype. Under this simulation scheme, Fig. 2 i also gives the actual associations, and all cells are associated with the phenotype.

The results of SCIPAC under the default resolution are shown in Fig. 2 j, k. Clearly, the associations SCIPAC identifies are highly consistent with the truth. Particularly, it successfully identifies the cells in the center as early-stage cells and most cells at the end of branches as last-stage cells. The results of SCIPAC under other resolutions are given in Additional file 1: Fig. S7 and S8, which are highly consistent. Scissor does not work with ordinal phenotypes; thus, no results are reported here.

Performance in real data

We consider four real datasets: a prostate cancer dataset, a breast cancer dataset, a lung cancer dataset, and a muscular dystrophy dataset. The bulk RNA-seq data of the three cancer datasets are obtained from the TCGA database, and that of the muscular dystrophy dataset is obtained from a published paper [ 28 ]. A detailed description of these datasets is given in Additional file 1. We will use these datasets to assess the performance of SCIPAC on different types of phenotypes. The cell type information (i.e., which cell belongs to which cell type) is available for the first three datasets, but we ignore this information so that we can make a fair comparison with Scissor, which cannot utilize this information.

Prostate cancer data with a binary phenotype

We use the single-cell expression of 8,700 cells from prostate-cancer tumors sequenced by [ 29 ]. The cell types of these cells are known and given in Fig. 3 a. The bulk data comprises 550 TCGA-PRAD (prostate adenocarcinoma) samples with phenotype (cancer vs. normal) information. Here the phenotype is cancer, and it is binary: present or absent.

UMAP visualization of the prostate cancer data, with a zoom-in view for the red-circled region (cell type MNP). a  True cell types. BE, HE, and CE stand for basal, hillock, club epithelial cells, LE-KLK3 and LE-KLK4 stand for luminal epithelial cells with high levels of kallikrein related peptidase 3 and 4, and MNP stands for mononuclear phagocytes. In the zoom-in view, the sub-types of MNP cells are given. b  Association strengths \(\Lambda\) given by SCIPAC under the default resolution. The cyan-circled cells are B cells, which are estimated by SCIPAC as negatively associated with cancer but estimated by Scissor as Scissor+ or null. c   p -values given by SCIPAC. The MNP cell type, which is red-circled in the plot, is estimated by SCIPAC to be strongly negatively associated with cancer but estimated by Scissor to be positively associated with cancer. d  Results given by Scissor under different \(\alpha\) values

Results from SCIPAC with the default resolution are shown in Fig. 3 b, c (results with other resolutions, given in Additional file 1: Fig. S9 and S10, are highly consistent with results here.) Compared with results from Scissor, shown in Fig. 3 d, results from SCIPAC again show three advantages. First, results from SCIPAC are richer and more comprehensive. SCIPAC gives estimated associations and the corresponding p -values, and the estimated associations are quantitative (shown in Fig. 3 b as different shades to the red or blue color) instead of discrete (shown in Fig. 3 d as a uniform shade to the red, blue, or light gray color). Second, SCIPAC’s results can be easier to interpret as the red and blue colors are more block-wise instead of scattered. Third, unlike Scissor, which produces multiple sets of results varying based on the parameter \(\alpha\) —a parameter without a default value or tuning guidance—typically, a single set of results from SCIPAC under its default settings suffices.

Comparing the results from our SCIPAC method with those from Scissor is non-trivial, as the latter’s outcomes are scattered and include multiple sets. We propose the following solutions to summarize the inferred association of a known cell type with the phenotype using a specific method (Scissor under a specific \(\alpha\) value, or SCIPAC with the default setting). We first calculate the proportion of cells in this cell type identified as Scissor+ (by Scissor at a specific \(\alpha\) value) or as significantly positively associated (by SCIPAC), denoted by \(p_{+}\) . We also calculate the proportion of all cells, encompassing any cell type, which are identified as Scissor+ or significantly positively associated, serving as the average background strength, denoted by \(p_{a}\) . Then, we compute the log odds ratio for this cell type to be positively associated with the phenotype compared to the background, represented as:

Similarly, the log odds ratio for the cell type to be negatively associated with the phenotype, \(\rho _-\) , is computed in a parallel manner.

For SCIPAC, a cell type is summarized as positively associated with the phenotype if \(\rho _+ \ge 1\) and \(\rho _- < 1\)  and negatively associated if \(\rho _- \ge 1\) and \(\rho _+ < 1\) . If neither condition is met, the association is inconclusive. For Scissor, we apply it under six different \(\alpha\) values: 0.01, 0.05, 0.10, 0.15, 0.20, and 0.25. A cell type is summarized as positively associated with the phenotype if \(\rho _+ \ge 1\) and \(\rho _- < 1\) in at least four of these \(\alpha\) values and negatively associated if \(\rho _- \ge 1\) and \(\rho _+ < 1\) in at least four \(\alpha\) values. If these criteria are not met, the association is deemed inconclusive. The above computation of log odds ratios and the determination of associations are performed only on cell types that each compose at least 1% of the cell population, to ensure adequate power.

For the prostate cancer data, the log odds ratios for each cell type using each method are presented in Tables S1 and S2. The final associations determined for each cell type are summarized in Table S3. In the last column of this table, we also indicate whether the conclusions drawn from SCIPAC and Scissor are consistent or not.

We find that SCIPAC’s results agree with Scissor on most cell types. However, there are three exceptions: mononuclear phagocytes (MNPs), B cells, and LE-KLK4.

MNPs are red-circled and zoomed in in each sub-figure of Fig. 3 . Most cells in this cell type are colored red in Fig. 3 d but colored dark blue in Fig. 3 b. In other words, while Scissor determines that this cell type is Scissor+, SCIPAC makes the opposite inference. Moreover, SCIPAC is confident about its judgment by giving small p -values, as shown in Fig. 3 c. To see which inference is closer to the biological fact is not easy, as biologically MNPs contain a number of sub-types that each have different functions [ 30 , 31 ]. Fortunately, this cell population has been studied in detail in the original paper that generated this dataset [ 29 ], and the sub-type information of each cell is provided there: this MNP population contains six sub-types, which are dendritic cells (DC), M1 macrophages (Mac1), metallothionein-expressing macrophages (Mac-MT), M2 macrophages (Mac2), proliferating macrophages (Mac-cycling), and monocytes (Mono), as shown in the zoom-in view of Fig. 3 a. Among these six sub-types, DC, Mac1, and Mac-MT are believed to inhibit cancer development and can serve as targets in cancer immunotherapy [ 29 ]; they compose more than 60% of all MNP cells in this dataset. SCIPAC makes the correct inference on this majority of MNP cells. Another cell type, Mac2, is reported to promote tumor development [ 32 ], but it only composes less than \(15\%\) of the MNPs. How the other two cell types, Mac-cycling and Mono, are associated with cancer is less studied. Overall, the results given by SCIPAC are more consistent with the current biological knowledge.

B cells are cyan-circled in Fig. 3 b. B cells are generally believed to have anti-tumor activity by producing tumor-reactive antibodies and forming tertiary lymphoid structures [ 29 , 33 ]. This means that B cells are likely to be negatively associated with cancer. SCIPAC successfully identifies this negative association, while Scissor fails.

LE-KLK4, a subtype of cancer cells, is thought to be positively associated with the tumor phenotype [ 29 ]. SCIPAC successfully identified this positive association, in contrast to Scissor, which failed to do so (in the figure, a proportion of LE-KLK4 cells are identified as Scissor+, especially under the smallest \(\alpha\) value; however, this proportion is not significantly higher than the background Scissor+ level under the majority of \(\alpha\) values).

In summary, across all three cell types, the results from SCIPAC appear to be more consistent with current biological knowledge. For more discussions regarding this dataset, refer to Additional file 1.

Breast cancer data with an ordinal phenotype

The scRNA-seq data for breast cancer are from [ 34 ], and we use the 19,311 cells from the five HER2+ tumor tissues. The true cell types are shown in Fig. 4 a. The bulk data include 1215 TCGA-BRCA samples with information on the cancer stage (I, II, III, or IV), which is treated as an ordinal phenotype.

UMAP visualization of the breast cancer data. a  True cell types. CAFs stand for cancer-associated fibroblasts, PB stands for plasmablasts and PVL stands for perivascular-like cells. b , c  Association strengths \(\Lambda\) and p -values given by SCIPAC under the default resolution. Cyan-circled are a group of T cells that are estimated by SCIPAC to be most significantly associated with the cancer stage in the negative direction, and orange-circled are a group of T cells that are estimated by SCIPAC to be significantly positively associated with the cancer stage. d  DE analysis of the cyan-circled T cells vs. all the other T cells. e  DE analysis of the cyan-circled T cells vs. all the other cells. f  Expression of CD8+ T cell marker genes in the cyan-circled cells and all the other cells. g  DE analysis of the orange-circled T cells vs. all the other cells. h  Expression of regulatory T cell marker genes in the orange-circled cells and all the other cells

Association strengths and p -values given by SCIPAC under the default resolution are shown in Fig. 4 b, c. Results under other resolutions are given in Additional file 1: Fig. S11 and S12, and again they are highly consistent with results under the default resolution. We do not present the results from Scissor, as Scissor does not take ordinal phenotypes.

In the SCIPAC results, cells that are most strongly and statistically significantly associated with the phenotype in the positive direction are the cancer-associated fibroblasts (CAFs). This finding agrees with the literature: CAFs contribute to therapy resistance and metastasis of cancer cells via the production of secreted factors and direct interaction with cancer cells [ 35 ], and they are also active players in breast cancer initiation and progression [ 36 , 37 , 38 , 39 ]. Another large group of cells identified as positively associated with the phenotype is the cancer epithelial cells. They are malignant cells in breast cancer tissues and are thus expected to be associated with severe cancer stages.

Of the cells identified as negatively associated with severe cancer stages, a large portion of T cells is the most noticeable. Biologically, T cells contain many sub-types, including CD4+, CD8+, regulatory T cells, and more, and their functions are diverse in the tumor microenvironment [ 40 ]. To explore SCIPAC’s discoveries, we compare T cells that are identified as most statistically significant, with p -values \(< 10^{-6}\) and circled in Fig. 4 d, with the other T cells. Differential expression (DE) analysis (details about DE analysis and other analyses are given in Additional file 1) identifies seven genes upregulated in these most significant T cells. Of these seven genes, at least five are supported by the literature: CCL4, XCL1, IFNG, and GZMB are associated with CD8+ T cell infiltration; they have been shown to have anti-tumor functions and are involved in cancer immunotherapy [ 41 , 42 , 43 ]. Also, IL2 has been shown to serve an important role in combination therapies for autoimmunity and cancer [ 44 ]. We also perform an enrichment analysis [ 45 ], in which a pathway called Myc stands out with a \(\textit{p}\text{-value}<10^{-7}\) , much smaller than all other pathways. Myc is downregulated in the T cells that are identified as most negatively associated with cancer stage progress. This agrees with current biological knowledge about this pathway: Myc is known to contribute to malignant cell transformation and tumor metastasis [ 46 , 47 , 48 ].

On the above, we have compared T cells that are most significantly associated with cancer stages in the negative direction with the other T cells using DE and pathway analysis, and the results could suggest that these cells are tumor-infiltrated CD8+ T cells with tumor-inhibition functions. To check this hypothesis, we perform DE analysis of these cells against all other cells (i.e., the other T cells and all the other cell types). The DE genes are shown in Fig. 4 e. It can be noted that CD8+ T cell marker genes such as CD8A, CD8B, and GZMK are upregulated. We further obtain CD8+ T cell marker genes from CellMarker [ 49 ] and check their expression, as illustrated in Fig. 4 f. Marker genes CD8A, CD8B, CD3D, GZMK, and CD7 show significantly higher expression in these T cells. This again supports our hypothesis that these cells are tumor-infiltrated CD8+ T cells that have anti-tumor functions.

Interestingly, not all T cells are identified as negatively associated with severe cancer stages; a group of T cells is identified as positively associated, as circled in Fig. 4 c. To explore the function of this group of T cells, we perform DE analysis of these T cells against the other T cells. The DE genes are shown in Fig. 4 g. Based on the literature, six out of eight over-expressed genes are associated with cancer development. The high expression of NUSAP1 gene is associated with poor patient overall survival, and this gene also serves as a prognostic factor in breast cancer [ 50 , 51 , 52 ]. Gene MKI67 has been treated as a candidate prognostic prediction for cancer proliferation [ 53 , 54 ]. The over-expression of RRM2 has been linked to higher proliferation and invasiveness of malignant cells [ 55 , 56 ], and the upregulation of RRM2 in breast cancer suggests it to be a possible prognostic indicator [ 57 , 58 , 59 , 60 , 61 , 62 ]. The high expression of UBE2C gene always occurs in cancers with a high degree of malignancy, low differentiation, and high metastatic tendency [ 63 ]. For gene TOP2A, it has been proposed that the HER2 amplification in HER2 breast cancers may be a direct result of the frequent co-amplification of TOP2A [ 64 , 65 , 66 ], and there is a high correlation between the high expressions of TOP2A and the oncogene HER2 [ 67 , 68 ]. Gene CENPF is a cell cycle-associated gene, and it has been identified as a marker of cell proliferation in breast cancers [ 69 ]. The over-expression of these genes strongly supports the correctness of the association identified by SCIPAC. To further validate this positive association, we perform DE analysis of these cells against all the other cells. We find that the top marker genes obtained from CellMarker [ 49 ] for the regulatory T cells, which are known to be immunosuppressive and promote cancer progression [ 70 ], are over-expressed with statistical significance, as shown in Fig. 4 h. This finding again provides strong evidence that the positive association identified by SCIPAC for this group of T cells is correct.

Lung cancer data with survival information

The scRNA-seq data for lung cancer are from [ 71 ], and we use two lung adenocarcinoma (LUAD) patients’ data with 29,888 cells. The true cell types are shown in Fig. 5 a. The bulk data consist of 576 TCGA-LUAD samples with survival status and time.

UMAP visualization of a–d  the lung cancer data and e–g  the muscular dystrophy data. a  True cell types. b , c  Association strengths \(\Lambda\) and p -values given by SCIPAC under the default resolution. d  Results given by Scissor under different \(\alpha\) values. e , f  Association strengths \(\Lambda\) and p -values given by SCIPAC under the default resolution. Circled are a group of cells that are identified by SCIPAC as significantly positively associated with the disease but identified by Scissor as null. g  Results given by Scissor under different \(\alpha\) values

Association strengths and p -values given by SCIPAC are given in Fig. 5 b, c (results under other resolutions are given in Additional file 1: Fig. S13 and S14). In Fig. 5 c, most cells with statistically significant associations are CD4+ T cells or B cells. These associations are negative, meaning that the abundance of these cells is associated with a reduced death rate, i.e., longer survival time. This agrees with the literature: CD4+ T cells primarily mediate anti-tumor immunity and are associated with favorable prognosis in lung cancer patients [ 72 , 73 , 74 ]; B cells also show anti-tumor functions in all stages of human lung cancer development and play an essential role in anti-tumor responses [ 75 , 76 ].

The results by Scissor under different \(\alpha\) values are shown in Fig. 5 d. The highly scattered Scissor+ and Scissor− cells make identifying and interpreting meaningful phenotype-associated cell groups difficult.

Muscular dystrophy data with a binary phenotype

This dataset contains cells from four facioscapulohumeral muscular dystrophy (FSHD) samples and two control samples [ 77 ]. We pool all the 7047 cells from these six samples together. The true cell types of these cells are unknown. The bulk data consists of 27 FSHD patients and eight controls from [ 28 ]. Here the phenotype is FSHD, and it is binary: present or absent.

The results of SCIPAC with the default resolution are given in Fig. 5 e, f. Results under other resolutions are highly similar (shown in Additional file 1: Fig. S15 and S16). For comparison, results given by Scissor under different \(\alpha\) values are presented in Fig. 5 g. The agreements between the results of SCIPAC and Scissor are clear. For example, both methods identify cells located at the top and lower left part of UMAP plots to be negatively associated with FSHD, and cells located at the center and right parts of UMAP plots to be positively associated. However, the discrepancies in their results are also evident. The most pronounced one is a large group of cells (circled in Fig. 5 f) that are identified by SCIPAC as significantly positively associated but are completely ignored by Scissor. Checking into this group of cells, we find that over 90% (424 out of 469) come from the FSHD patients, and less than 10% come from the control samples. However, cells from FSHD patients only compose 73% (5133) of all the 7047 cells. This statistically significant ( p -value \(<10^{-15}\) , Fisher’s exact test) over-representation (odds ratio = 3.51) suggests that the positive association identified SCIPAC is likely to be correct.

SCIPAC is computationally highly efficient. On an 8-core machine with 2.50 GHz CPU and 16 GB RAM, SCIPAC takes 7, 24, and 2 s to finish all the computation and give the estimated association strengths and p -values on the prostate cancer, lung cancer, and muscular dystrophy datasets, respectively. As a reference, Scissor takes 314, 539, and 171 seconds, respectively.

SCIPAC works with various phenotype types, including binary, continuous, survival, and ordinal. It can easily accommodate other types by using a proper regression model with a systematic component in the form of Eq. 3 (see the “ Methods ” section). For example, a Poisson or negative binomial log-linear model can be used if the phenotype is a count (i.e., non-negative integer).

In SCIPAC’s definition of association, a cell type is associated with the phenotype if increasing the proportion of this cell type leads to a change of probability of the phenotype occurring. The strength of association represents the extent of the increase or decrease in this probability. In the case of binary-response, this change is measured by the log odds ratio. For example, if the association strength of cell type A is twice that of cell type B, increasing cell type A by a certain proportion leads to twice the amount of change in the log odds ratio of having the phenotype compared to increasing cell type B by the same proportion. The association strength under other types of phenotypes can be interpreted similarly, with the major difference lying in the measure of change in probability. For quantitative, ordinal, and survival outcomes, the difference in the quantitative outcome, log odds ratio of the right-tail probability, and log hazard ratio respectively are used. Despite the differences in the exact form of the association strength under different types of phenotypes, the underlying concept remains the same: a larger (absolute value of) association strength indicates that the same increase/decrease in a cell type leads to a larger change in the occurrence of the phenotype.

As SCIPAC utilizes both bulk RNA-seq data with phenotype and single-cell RNA-seq data, the estimated associations for the cells are influenced by the choice of the bulk data. Although different bulk data can yield varying estimations of the association for the same single cells, the estimated associations appear to be reasonably robust even when minor changes are made to the bulk data. See Additional file 1 for further discussions.

When using the Louvain algorithm in the Seurat package to cluster cells, SCIPAC’s default resolution is 2.0, larger than the default setting of Seurat. This allows for the identification of potential subtypes within the major cell type and enables the estimation of individual association strengths. Consequently, a more detailed and comprehensive description of the association between single cells and the phenotype can be obtained by SCIPAC.

When applying SCIPAC to real datasets, we made a deliberate choice to disregard the cell annotation provided by the original publications and instead relied on the inferred cell clusters produced by the Louvain algorithm. We made this decision for several reasons. Firstly, we aimed to ensure a fair comparison with Scissor, as it does not utilize cell-type annotations. Secondly, the original annotation might not be sufficiently comprehensive or detailed. Presumed cell types could potentially encompass multiple subtypes, each of which may exhibit distinct associations with the phenotype under investigation. In such cases, employing the Louvain algorithm with a relatively high resolution, which is the default setting in SCIPAC, enables us to differentiate between these subtypes and allows SCIPAC to assign varying association strengths to each subtype.

SCIPAC fits the regression model using the elastic net, a machine-learning algorithm that maximizes a penalized version of the likelihood. The elastic net can be replaced by other penalized estimates of regression models, such as SCAD [ 78 ], without altering the rest of the SCIPAC algorithm. The combination of a regression model and a penalized estimation algorithm such as the elastic net has shown comparable or higher prediction power than other sophisticated methods such as random forests, boosting, or neural networks in numerous applications, especially for gene expression data [ 79 ]. However, there can still be datasets where other models have higher prediction power. It will be future work to incorporate these models into SCIPAC.

The use of metacells is becoming an efficient way to handle large single-cell datasets [ 80 , 81 , 82 , 83 ]. Conceptually, SCIPAC can incorporate metacells and their representatives as an alternative to its default setting of using cell clusters/types and their centroids. We have explored this aspect using metacells provided by SEACells [ 81 ]. Details are given in Additional file 1. Our comparative analysis reveals that combining SCIPAC with SEACells results in significantly reduced performance compared to using SCIPAC directly on original single-cell data. The primary reason for this appears to be the subpar performance of SEACells in cell grouping, especially when contrasted with the Louvain algorithm. Given these findings, we do not suggest using metacells provided by SEACells for SCIPAC applications in the current stage.

Conclusions

SCIPAC is a novel algorithm for studying the associations between cells and phenotypes. Compared to the previous algorithm, SCIPAC gives a much more detailed and comprehensive description of the associations by enabling a quantitative estimation of the association strength and by providing a quality control—the p -value. Underlying SCIPAC are a general statistical model that accommodates virtually all types of phenotypes, including ordinal (and potentially count) phenotypes that have never been considered before, and a concise and closed-form mathematical formula that quantifies the association, which minimizes the computational load. The mathematical conciseness also largely frees SCIPAC from parameter tuning. The only parameter (i.e., the resolution) barely changes the results given by SCIPAC. Overall, compared with its predecessor, SCIPAC represents a substantially more capable software by being much more informative, versatile, robust, and user-friendly.

The improvement in accuracy is also remarkable. In simulated data, SCIPAC achieves high power and low false positives, which is evident from the UMAP plot, F1 score, and FSC score. In real data, SCIPAC gives results that are consistent with current biological knowledge for cell types whose functions are well understood. For cell types whose functions are less studied or more multifaceted, SCIPAC gives support to certain biological hypotheses or helps identify/discover cell sub-types.

SCIPAC’s identification of cell-phenotype associations closely follows its definition of association: when increasing the fraction of a cell type increases (or decreases) the probability for a phenotype to be present, this cell type is positively (or negatively) associated with the phenotype.

The increase of the fraction of a cell type

For a bulk sample, let vector \(\varvec{G} \in \mathbb {R}^p\) be its expression profile, that is, its expression on the p genes. Suppose there are K cell types in the tissue, and let \(\varvec{g}_{k}\) be the representative expression of the k ’th cell type. Usually, people assume that \(\varvec{G}\) can be decomposed by

where \(\gamma _{k}\) is the proportion of cell type k in the bulk tissue, with \(\sum _{k = 1}^{K}\gamma _{k} = 1\) . This equation links the bulk and single-cell expression data.

Now consider increasing cells from cell type k by \(\Delta \gamma\) proportion of the original number of cells. Then, the new proportion of cell type k becomes \(\frac{\gamma _{k} + \Delta \gamma }{1 + \Delta \gamma }\) , and the new proportion of cell type \(j \ne k\) becomes \(\frac{\gamma _{j}}{1 + \Delta \gamma }\)  (note that the new proportions of all cell types should still add up to 1). Thus, the bulk expression profile with the increase of cell type k becomes

Plugging Eq. 1 , we get

Interestingly, this expression of \(\varvec{G}^*\) does not include \(\gamma _{1}, \ldots , \gamma _{K}\) . This means that there is no need actually to compute \(\gamma _{1}, \ldots , \gamma _{K}\) in Eq. 1 , which could otherwise be done using a cell-type-decomposition software, but an accurate and robust decomposition is non-trivial [ 84 , 85 , 86 ]. See Additional file 1 for a more in-depth discussion on the connections of SCIPAC with decomposition/deconvolution.

The change in chance of a phenotype

In this section, we consider how the increase in the fraction of a cell type will change the chance for a binary phenotype such as cancer to occur. Other types of phenotypes will be considered in the next section.

Let \(\pi (\varvec{G})\) be the chance of an individual with gene expression profile \(\varvec{G}\) for this phenotype to occur. We assume a logistic regression model to describe the relationship between \(\pi (\varvec{G})\) and \(\varvec{G}\) :

here the left-hand side is the log odds of \(\pi (\varvec{G})\) , \(\beta _{0}\) is the intercept, and \(\varvec{\beta }\) is a length- p vector of coefficients. In the section after the next, we will describe how we obtain \(\beta _{0}\) and \(\varvec{\beta }\) from the data.

When increasing cells from cell type k by \(\Delta \gamma\) , \(\varvec{G}\) becomes \(\varvec{G}^*\) in Eq. 3 . Plugging Eq. 2 , we get

We further take the difference between Eqs. 4 and 3 and get

The left-hand side of this equation is the log odds ratio (i.e., the change of log odds). On the right-hand side, \(\frac{\Delta \gamma }{1 + \Delta \gamma }\) is an increasing function with respect to \(\Delta \gamma\) , and \(\varvec{\beta }^T(\varvec{g}_{k} - \varvec{G})\) is independent of \(\Delta \gamma\) . This indicates that given any specific \(\Delta \gamma\) , the log odds ratio under over-representation of cell type k is proportional to

\(\lambda _k\) describes the strength of the effect of increasing cell type k to a bulk sample with expression profile \(\varvec{G}\) . Given the presence of numerous bulk samples, employing multiple \(\lambda _k\) ’s could be cumbersome and obscure the overall effect of a particular cell type. To concisely summarize the association of cell type k , we propose averaging their effects. The average effect on all bulk samples can be obtained by

where \(\bar{\varvec{G}}\) is the average expression profile of all bulk samples.

\(\Lambda _k\) gives an overall impression of how strong the effect is when cell type k over-represents to the probability for the phenotype to be present. Its sign represents the direction of the change: a positive value means an increase in probability, and a negative value means a decrease in probability. Its absolute value represents the strength of the effect. In SCIPAC, we call \(\Lambda _k\) the association strength of cell type k and the phenotype.

Note that this derivation does not involve likelihood, although the computation of \(\varvec{\beta }\) does. Here, it serves more as a definitional approach.

Definition of the association strength for other types of phenotype

Our definition of \(\Lambda _k\) relies on vector \(\varvec{\beta }\) . In the case of a binary phenotype, \(\varvec{\beta }\) are the coefficients of a logistic regression that describes a linear relationship between the expression profile and the log odds of having the phenotype, as shown in Eq. 3 . For other types of phenotype, \(\varvec{\beta }\) can be defined/computed similarly.

For a quantitative (i.e., continuous) phenotype, an ordinary linear regression can be used, and the left-hand side of Eq. 3 is changed to the quantitative value of the phenotype.

For a survival phenotype, a Cox proportional hazards model can be used, and the left-hand side of Eq. 3 is changed to the log hazard ratio.

For an ordinal phenotype, we use a proportional odds model

where \(j \in \{1, 2, ..., (J - 1)\}\) and J is the number of ordinal levels. It should be noted that here we use the right-tail probability \(\Pr (Y_{i} \ge j + 1 | X)\) instead of the commonly used cumulative probability (left-tail probability) \(\Pr (Y_{i} \le j | X)\) . Such a change makes the interpretation consistent with other types of phenotypes: in our model, a larger value on the right-hand side indicates a larger chance for \(Y_{i}\) to have a higher level, which in turn guarantees that the sign of the association strength defined according to this \(\varvec{\beta }\) has the usual meaning: a positive \(\Lambda _k\) value means a positive association with the phenotype-using the cancer stage as an example. A positive \(\Lambda _k\) means the over-representation of cell type k increases the chance of a higher cancer stage. In contrast, using the commonly used cumulative probability leads to a counter-intuitive, reversed interpretation.

Computation of the association strength in practice

In practice, \(\varvec{\beta }\) in Eq. 3 needs to be learned from the bulk data. By default, SCIPAC uses the elastic net, a popular and powerful penalized regression method:

In this model, \(l(\beta _{0}, \varvec{\beta })\) is a log-likelihood of the linear model (i.e., logistic regression for a binary phenotype, ordinary linear regression for a quantitative phenotype, Cox proportional odds model for a survival phenotype, and proportional odds model for an ordinal phenotype). \(\alpha\) is a number between 0 and 1, denoting a combination of \(\ell _1\) and \(\ell _2\) penalties, and \(\lambda\) is the penalty strength. SCIPAC fixes \(\alpha\) to be 0.4 (see Additional file 1 for discussions on this choice) and uses 10-fold cross-validation to decide \(\lambda\) automatically. This way, they do not become hyperparameters.

In SCIPAC, the fitting and cross-validation of the elastic net are done by calling the ordinalNet [ 87 ] R package for the ordinal phenotype and by calling the glmnet R package [ 88 , 89 , 90 , 91 ] for other types of phenotypes.

The computation of the association strength, as defined by Eq. 7 , does not only require \(\varvec{\beta }\) , but also \(\varvec{g}_k\) and \(\bar{\varvec{G}}\) . \(\bar{\varvec{G}}\) is simply the average expression profile of all bulk samples. On the other hand, \(\varvec{g}_k\) requires knowing the cell type of each cell. By default, SCIPAC does not assume this information to be given, and it uses the Louvain clustering implemented in the Seurat [ 24 , 25 ] R package to infer it. This clustering algorithm has one tuning parameter called “resolution.” SCIPAC sets its default value as 2.0, and the user can use other values. With the inferred or given cell types, \(\varvec{g}_k\) is computed as the centroid (i.e., the mean expression profile) of cells in cluster k .

Given \(\varvec{\beta }\) , \(\bar{\varvec{G}}\) , and \(\varvec{g}_k\) , the association strength can be computed using Eq. 7 . Knowing the association strength for each cell type and the cell-type label for each cell, we also know the association strength for every single cell. In practice, we standardize the association strengths for all cells. That is, we compute the mean and standard deviation of the association strengths of all cells and use them to centralize and scale the association strength, respectively. We have found such standardization makes SCIPAC more robust to the possible unbalance in sample size of bulk data in different phenotype groups.

Computation of the p -value

SCIPAC uses non-parametric bootstrap [ 92 ] to compute the standard deviation and hence the p -value of the association. Fifty bootstrap samples, which are believed to be enough to compute the standard error of most statistics [ 93 ], are generated for the bulk expression data, and each is used to compute (standardized) \(\Lambda\) values for all the cells. For cell i , let its original \(\Lambda\) values be \(\Lambda _i\) , and the bootstrapped values be \(\Lambda _i^{(1)}, \ldots , \Lambda _i^{(50)}\) . A z -score is then computed using

and then the p -value is computed according to the cumulative distribution function of the standard Gaussian distribution. See Additional file 1 for more discussions on the calculation of p -value.

Availability of data and materials

The simulated datasets [ 94 ] under three schemes are available at Zenodo with DOI 10.5281/zenodo.11013320 [ 95 ]. The SCIPAC package is available at GitHub website https://github.com/RavenGan/SCIPAC under the MIT license [ 96 ]. The source code of SCIPAC is also deposited at Zenodo with DOI 10.5281/zenodo.11013696 [ 97 ]. A vignette of the R package is available on the GitHub page and in the Additional file 2. The prostate cancer scRNA-seq data is obtained from the Prostate Cell Atlas https://www.prostatecellatlas.org [ 29 ]; the scRNA-seq data for the breast cancer are from the Gene Expression Omnibus (GEO) under accession number GSE176078 [ 34 , 98 ]; the scRNA-seq data for the lung cancer are from E-MTAB-6149 [ 99 ] and E-MTAB-6653 [ 71 , 100 ]; the scRNA-seq data for facioscapulohumeral muscular dystrophy data are from the GEO under accession number GSE122873 [ 101 ]. The bulk RNA-seq data are obtained from the TCGA database via TCGAbiolinks (ver. 2.25.2) R package [ 102 ]. More details about the simulated and real scRNA-seq and bulk RNA-seq data can be found in the Additional file 1.

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Veronique van den Berghe was the primary editor of this article and managed its editorial process and peer review in collaboration with the rest of the editorial team.

This work is supported by the National Institutes of Health (R01CA280097 to X.L. and J.L, R01CA252878 to J.L.) and the DOD BCRP Breakthrough Award, Level 2 (W81XWH2110432 to J.L.).

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Dailin Gan & Jun Li

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J.L. conceived and supervised the study. J.L. and D.G. proposed the methods. D.G. implemented the methods and analyzed the data. D.G. and J.L. drafted the paper. D.G., Y.Z., X.L., and J.L. interpreted the results and revised the paper.

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Gan, D., Zhu, Y., Lu, X. et al. SCIPAC: quantitative estimation of cell-phenotype associations. Genome Biol 25 , 119 (2024). https://doi.org/10.1186/s13059-024-03263-1

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Artificial intelligence in strategy

Can machines automate strategy development? The short answer is no. However, there are numerous aspects of strategists’ work where AI and advanced analytics tools can already bring enormous value. Yuval Atsmon is a senior partner who leads the new McKinsey Center for Strategy Innovation, which studies ways new technologies can augment the timeless principles of strategy. In this episode of the Inside the Strategy Room podcast, he explains how artificial intelligence is already transforming strategy and what’s on the horizon. This is an edited transcript of the discussion. For more conversations on the strategy issues that matter, follow the series on your preferred podcast platform .

Joanna Pachner: What does artificial intelligence mean in the context of strategy?

Yuval Atsmon: When people talk about artificial intelligence, they include everything to do with analytics, automation, and data analysis. Marvin Minsky, the pioneer of artificial intelligence research in the 1960s, talked about AI as a “suitcase word”—a term into which you can stuff whatever you want—and that still seems to be the case. We are comfortable with that because we think companies should use all the capabilities of more traditional analysis while increasing automation in strategy that can free up management or analyst time and, gradually, introducing tools that can augment human thinking.

Joanna Pachner: AI has been embraced by many business functions, but strategy seems to be largely immune to its charms. Why do you think that is?

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Yuval Atsmon: You’re right about the limited adoption. Only 7 percent of respondents to our survey about the use of AI say they use it in strategy or even financial planning, whereas in areas like marketing, supply chain, and service operations, it’s 25 or 30 percent. One reason adoption is lagging is that strategy is one of the most integrative conceptual practices. When executives think about strategy automation, many are looking too far ahead—at AI capabilities that would decide, in place of the business leader, what the right strategy is. They are missing opportunities to use AI in the building blocks of strategy that could significantly improve outcomes.

I like to use the analogy to virtual assistants. Many of us use Alexa or Siri but very few people use these tools to do more than dictate a text message or shut off the lights. We don’t feel comfortable with the technology’s ability to understand the context in more sophisticated applications. AI in strategy is similar: it’s hard for AI to know everything an executive knows, but it can help executives with certain tasks.

When executives think about strategy automation, many are looking too far ahead—at AI deciding the right strategy. They are missing opportunities to use AI in the building blocks of strategy.

Joanna Pachner: What kind of tasks can AI help strategists execute today?

Yuval Atsmon: We talk about six stages of AI development. The earliest is simple analytics, which we refer to as descriptive intelligence. Companies use dashboards for competitive analysis or to study performance in different parts of the business that are automatically updated. Some have interactive capabilities for refinement and testing.

The second level is diagnostic intelligence, which is the ability to look backward at the business and understand root causes and drivers of performance. The level after that is predictive intelligence: being able to anticipate certain scenarios or options and the value of things in the future based on momentum from the past as well as signals picked in the market. Both diagnostics and prediction are areas that AI can greatly improve today. The tools can augment executives’ analysis and become areas where you develop capabilities. For example, on diagnostic intelligence, you can organize your portfolio into segments to understand granularly where performance is coming from and do it in a much more continuous way than analysts could. You can try 20 different ways in an hour versus deploying one hundred analysts to tackle the problem.

Predictive AI is both more difficult and more risky. Executives shouldn’t fully rely on predictive AI, but it provides another systematic viewpoint in the room. Because strategic decisions have significant consequences, a key consideration is to use AI transparently in the sense of understanding why it is making a certain prediction and what extrapolations it is making from which information. You can then assess if you trust the prediction or not. You can even use AI to track the evolution of the assumptions for that prediction.

Those are the levels available today. The next three levels will take time to develop. There are some early examples of AI advising actions for executives’ consideration that would be value-creating based on the analysis. From there, you go to delegating certain decision authority to AI, with constraints and supervision. Eventually, there is the point where fully autonomous AI analyzes and decides with no human interaction.

Because strategic decisions have significant consequences, you need to understand why AI is making a certain prediction and what extrapolations it’s making from which information.

Joanna Pachner: What kind of businesses or industries could gain the greatest benefits from embracing AI at its current level of sophistication?

Yuval Atsmon: Every business probably has some opportunity to use AI more than it does today. The first thing to look at is the availability of data. Do you have performance data that can be organized in a systematic way? Companies that have deep data on their portfolios down to business line, SKU, inventory, and raw ingredients have the biggest opportunities to use machines to gain granular insights that humans could not.

Companies whose strategies rely on a few big decisions with limited data would get less from AI. Likewise, those facing a lot of volatility and vulnerability to external events would benefit less than companies with controlled and systematic portfolios, although they could deploy AI to better predict those external events and identify what they can and cannot control.

Third, the velocity of decisions matters. Most companies develop strategies every three to five years, which then become annual budgets. If you think about strategy in that way, the role of AI is relatively limited other than potentially accelerating analyses that are inputs into the strategy. However, some companies regularly revisit big decisions they made based on assumptions about the world that may have since changed, affecting the projected ROI of initiatives. Such shifts would affect how you deploy talent and executive time, how you spend money and focus sales efforts, and AI can be valuable in guiding that. The value of AI is even bigger when you can make decisions close to the time of deploying resources, because AI can signal that your previous assumptions have changed from when you made your plan.

Joanna Pachner: Can you provide any examples of companies employing AI to address specific strategic challenges?

Yuval Atsmon: Some of the most innovative users of AI, not coincidentally, are AI- and digital-native companies. Some of these companies have seen massive benefits from AI and have increased its usage in other areas of the business. One mobility player adjusts its financial planning based on pricing patterns it observes in the market. Its business has relatively high flexibility to demand but less so to supply, so the company uses AI to continuously signal back when pricing dynamics are trending in a way that would affect profitability or where demand is rising. This allows the company to quickly react to create more capacity because its profitability is highly sensitive to keeping demand and supply in equilibrium.

Joanna Pachner: Given how quickly things change today, doesn’t AI seem to be more a tactical than a strategic tool, providing time-sensitive input on isolated elements of strategy?

Yuval Atsmon: It’s interesting that you make the distinction between strategic and tactical. Of course, every decision can be broken down into smaller ones, and where AI can be affordably used in strategy today is for building blocks of the strategy. It might feel tactical, but it can make a massive difference. One of the world’s leading investment firms, for example, has started to use AI to scan for certain patterns rather than scanning individual companies directly. AI looks for consumer mobile usage that suggests a company’s technology is catching on quickly, giving the firm an opportunity to invest in that company before others do. That created a significant strategic edge for them, even though the tool itself may be relatively tactical.

Joanna Pachner: McKinsey has written a lot about cognitive biases  and social dynamics that can skew decision making. Can AI help with these challenges?

Yuval Atsmon: When we talk to executives about using AI in strategy development, the first reaction we get is, “Those are really big decisions; what if AI gets them wrong?” The first answer is that humans also get them wrong—a lot. [Amos] Tversky, [Daniel] Kahneman, and others have proven that some of those errors are systemic, observable, and predictable. The first thing AI can do is spot situations likely to give rise to biases. For example, imagine that AI is listening in on a strategy session where the CEO proposes something and everyone says “Aye” without debate and discussion. AI could inform the room, “We might have a sunflower bias here,” which could trigger more conversation and remind the CEO that it’s in their own interest to encourage some devil’s advocacy.

We also often see confirmation bias, where people focus their analysis on proving the wisdom of what they already want to do, as opposed to looking for a fact-based reality. Just having AI perform a default analysis that doesn’t aim to satisfy the boss is useful, and the team can then try to understand why that is different than the management hypothesis, triggering a much richer debate.

In terms of social dynamics, agency problems can create conflicts of interest. Every business unit [BU] leader thinks that their BU should get the most resources and will deliver the most value, or at least they feel they should advocate for their business. AI provides a neutral way based on systematic data to manage those debates. It’s also useful for executives with decision authority, since we all know that short-term pressures and the need to make the quarterly and annual numbers lead people to make different decisions on the 31st of December than they do on January 1st or October 1st. Like the story of Ulysses and the sirens, you can use AI to remind you that you wanted something different three months earlier. The CEO still decides; AI can just provide that extra nudge.

Joanna Pachner: It’s like you have Spock next to you, who is dispassionate and purely analytical.

Yuval Atsmon: That is not a bad analogy—for Star Trek fans anyway.

Joanna Pachner: Do you have a favorite application of AI in strategy?

Yuval Atsmon: I have worked a lot on resource allocation, and one of the challenges, which we call the hockey stick phenomenon, is that executives are always overly optimistic about what will happen. They know that resource allocation will inevitably be defined by what you believe about the future, not necessarily by past performance. AI can provide an objective prediction of performance starting from a default momentum case: based on everything that happened in the past and some indicators about the future, what is the forecast of performance if we do nothing? This is before we say, “But I will hire these people and develop this new product and improve my marketing”— things that every executive thinks will help them overdeliver relative to the past. The neutral momentum case, which AI can calculate in a cold, Spock-like manner, can change the dynamics of the resource allocation discussion. It’s a form of predictive intelligence accessible today and while it’s not meant to be definitive, it provides a basis for better decisions.

Joanna Pachner: Do you see access to technology talent as one of the obstacles to the adoption of AI in strategy, especially at large companies?

Yuval Atsmon: I would make a distinction. If you mean machine-learning and data science talent or software engineers who build the digital tools, they are definitely not easy to get. However, companies can increasingly use platforms that provide access to AI tools and require less from individual companies. Also, this domain of strategy is exciting—it’s cutting-edge, so it’s probably easier to get technology talent for that than it might be for manufacturing work.

The bigger challenge, ironically, is finding strategists or people with business expertise to contribute to the effort. You will not solve strategy problems with AI without the involvement of people who understand the customer experience and what you are trying to achieve. Those who know best, like senior executives, don’t have time to be product managers for the AI team. An even bigger constraint is that, in some cases, you are asking people to get involved in an initiative that may make their jobs less important. There could be plenty of opportunities for incorpo­rating AI into existing jobs, but it’s something companies need to reflect on. The best approach may be to create a digital factory where a different team tests and builds AI applications, with oversight from senior stakeholders.

The big challenge is finding strategists to contribute to the AI effort. You are asking people to get involved in an initiative that may make their jobs less important.

Joanna Pachner: Do you think this worry about job security and the potential that AI will automate strategy is realistic?

Yuval Atsmon: The question of whether AI will replace human judgment and put humanity out of its job is a big one that I would leave for other experts.

The pertinent question is shorter-term automation. Because of its complexity, strategy would be one of the later domains to be affected by automation, but we are seeing it in many other domains. However, the trend for more than two hundred years has been that automation creates new jobs, although ones requiring different skills. That doesn’t take away the fear some people have of a machine exposing their mistakes or doing their job better than they do it.

Joanna Pachner: We recently published an article about strategic courage in an age of volatility  that talked about three types of edge business leaders need to develop. One of them is an edge in insights. Do you think AI has a role to play in furnishing a proprietary insight edge?

Yuval Atsmon: One of the challenges most strategists face is the overwhelming complexity of the world we operate in—the number of unknowns, the information overload. At one level, it may seem that AI will provide another layer of complexity. In reality, it can be a sharp knife that cuts through some of the clutter. The question to ask is, Can AI simplify my life by giving me sharper, more timely insights more easily?

Joanna Pachner: You have been working in strategy for a long time. What sparked your interest in exploring this intersection of strategy and new technology?

Yuval Atsmon: I have always been intrigued by things at the boundaries of what seems possible. Science fiction writer Arthur C. Clarke’s second law is that to discover the limits of the possible, you have to venture a little past them into the impossible, and I find that particularly alluring in this arena.

AI in strategy is in very nascent stages but could be very consequential for companies and for the profession. For a top executive, strategic decisions are the biggest way to influence the business, other than maybe building the top team, and it is amazing how little technology is leveraged in that process today. It’s conceivable that competitive advantage will increasingly rest in having executives who know how to apply AI well. In some domains, like investment, that is already happening, and the difference in returns can be staggering. I find helping companies be part of that evolution very exciting.

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