It is then possible to analyze whether features are modified in a treated sample of cells compared to controls. The collection of features for a cell is called a profile (sometimes described as a morphological profile or image-based profile), and typically a thousand or more features are measured per cell.
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Diverse stains are used (as in the Cell Painting assay, which stains eight cell components Bray et al., 2016 Cimini et al., 2022) and then image analysis software segments the cells and measures all possible morphological features from single cells. By contrast, in image-based profiling, the aim is to let the cells speak for themselves. In a typical quantitative microscopy experiment, biologists select fluorescent biomarkers (such as antibodies or dyes for specific proteins or cell compartments) and measure only the features they hypothesize will be perturbed in the experiment.
Current Protocols published by Wiley Periodicals LLC.īasic Protocol 1: Exploratory analysis of profile similarities and driving featuresīasic Protocol 2: Image and single-cell visualization following profile interpretationĪutomated microscopy allows biologists to acquire thousands of images from cells perturbed with drugs, small interfering RNA (siRNA), CRISPR-Cas9, and more. Together, these two tutorials help researchers interpret image-based data to speed up research.
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In the second protocol, we show how to interactively explore images together with the numerical data, and we provide scripts to create visualizations of representative single cells and image sites to understand how changes in features are reflected in the images. The protocol includes steps to examine feature-driving differences between samples and to visualize correlations between features and treatments to create interpretable heatmaps using the open-source web tool Morpheus. In the first protocol, we examine the similarity among perturbed cell samples using data from compounds that cluster by their mechanisms of action.
Here, we provide two complementary protocols to help explore and interpret data from image-based profiling experiments. Image-based profiling quantitatively assesses the effects of perturbations on cells by capturing a breadth of changes via microscopy.
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