Cell Painting

Nucleus
Dye: Hoechst 33342

Mitochondria
Dye: MitoTracker Deep Red

Endoplasmic reticulum
Dye: Concanavalin A/Alexa Fluor 488 conjugate

Nucleoli, cytoplasmic RNA
Dye: SYT0 14 green fluorescent nucleic acid stain

F-actin cytoskeleton, Golgi, plasma membrane
Dye: Phalloidin/Alexa Fluor 568 conjugate, wheatgerm agglutinin/Alexa Fluor 555 conjugate
 

Composite image consisting of actin, ER (endoplasmic reticulum), and the nuclei.

Cell Painting for phenotypic profiling

The phenotypic profile of a cell reveals the specific biological state of a cell. More specifically, it can be used to interrogate biological perturbations because the cellular morphology is influenced by factors such as metabolism, genetic and epigenetic state of the cell, and environmental cues. In addition, it can be used to characterize healthy cells from diseased cells. Because a phenotypic profile is an aggregation of a large number of measurements, it is more sensitive to deviations or changes to those features extracted from Cell Painting assay. In other words, a phenotypic profile can capture certain characteristics of the cells that may not be obvious to the naked eye.

In an interview with the Science Explorer, Angeline Lim, PhD., explained, “I like to think of Cell Painting or phenotypic profiling as analogous to facial recognition. It's a little bit like how Facebook or iPhoto tags people. Once you have a photograph and you tag it, the software in the background extracts information about the photo and calculates a profile. When another photo pops up, the software compares its profile to that of the previous photo to see if they are the same or different. This is basically how Cell Painting works. With Cell Painting, we expect the image analysis part to cluster like cells from unlike cells, or diseased cells from healthy cells. This potentially has many applications, especially in drug discovery.”

Examples of treated cells and phenotypic changes

Here is an example in which treated cells show obvious changes in their phenotypes—only three of the five wavelengths are shown in this composite: nuclei in blue, endoplasmic reticulum in green, actin & Golgi in red.

 

Control, untreated cells.

Cells treated with rotenone, a toxin frequently used in insecticides. It is also known to inhibit mitochondrial ATP production and has been shown to have anticancer activity in various cancer cells.

Cells treated with chloroquine. Chloroquine was first developed for malaria treatment.

General workflow for Cell Painting assay

Learn more about Cell Painting

Cell Painting is emerging as a valuable tool with its many important potential applications particularly in drug discovery. Learn how to optimize high-content imaging capabilities and perform multiplexed assays using preconfigured or custom modules from MetaXpress software to provide a data rich, phenotypic profile.