Cell Painting is a high-throughput, phenotypic profiling assay that uses fluorescent cytochemistry o visualize a variety of organelles and high-content imaging to derive a large number of morphological features at the single cell level. Here, we used the Cell Painting assay to characterize the phenotypic effects of sixteen phenotypic reference chemicals in concentration- response screening mode across six biologically diverse human-derived cell lines (U-2 OS, MCF7, HepG2, A549, HTB-9, ARPE-19). All cell lines were labeled using the same cytochemistry protocol and the same set of phenotypic features were calculated. We found it necessary to optimize image acquisition settings and cell segmentation parameters for each cell type but did not adjust the cytochemistry protocol. For some reference chemicals, similar subsets of phenotypic features corresponding to a particular organelle were associated with the highest effect magnitudes in each affected cell type. Overall, for certain chemicals the Cell Painting assay yielded qualitatively similar biological activity profiles across a group of diverse, morphologically distinct human-derived cell lines without the requirement for cell-type specific optimization of cytochemistry protocols.

This dataset is associated with the following publication: Willis, C., J. Nyffeler, and J. Harrill. Phenotypic Profiling of Reference Chemicals Across Biologically Diverse Cell Types Using the Cell Painting Assay. SLAS Discovery. SAGE Publications, THOUSAND OAKS, CA, USA, 25(7): 755-769, (2020).