REMEDi4ALL - Standard Operating Procedure - Cell Painting assay for mechanism of action (MoA) studies

Cell Painting (CP) is a high-content imaging and analysis technique used to capture and quantify phenotypic changes in cells induced by various perturbations, such as small molecules, as well as other modalities such as PROTACS, biologics, and genetic perturbation such as overexpression or depletion of genes. The method involves staining cells with a combination of fluorescent dyes that stain different cellular components, including the nucleus, cytoskeleton, mitochondria, endoplasmic reticulum, nucleoli, cytoplasmic RNA and Golgi apparatus. By analysing the resulting images, profiles of each condition or treatment are generated, which are informative of the cellular responses. These profiles facilitate the identification of compound mechanisms of action, potential drug targets, and off-target effects. CP involves advanced image analysis to extract and analyze thousands of morphological features, and it requires powerful data processing of the millions to billions of data points generated.

This SOP describes the process of conducting CP experiments for mechanism of action studies.