What is flow cytometry?

Flow cytometry is a widely used, laser-based method for analyzing the expression of cell surface and intracellular molecules. There are many applications of flow cytometry in research and diagnostics, including simultaneous, multiparameter analysis of single cells and characterizing and defining different cell types in heterogeneous cell populations. Flow cytometry assays can also assess the purity of isolated subpopulations, analyze cell size and volume, and sort different cell populations, known as fluorescence-activated cell sorting (FACS)¹.

A flow cytometry test measures fluorescence intensity produced by fluorescently labeled antibodies specific to proteins on or in cells or ligands that bind to specific cell-associated molecules, such as propidium iodide binding to DNA.

The staining procedure involves making a single-cell suspension from cell culture or tissue samples. Then, the cells are incubated in tubes or microtiter plates with unlabeled or fluorophore-labeled antibodies and analyzed on the flow cytometer.

Multicolor flow cytometry takes this further by analyzing multiple parameters on thousands of single cells or other particles in seconds^2,3. In multicolor flow cytometry, fluorescent markers are used to characterize and define different cell types of interest in heterogeneous cell populations, assess the purity of isolated subpopulations, and analyze cell size and shape.

In this guide, we’ll take you through flow cytometry basics, principles, protocols, and analysis to give you an advanced understanding of how flow cytometry works, when it is valuable, and how to do it. You will also find everything you need to quickly and easily set up your multicolor flow cytometry experiment, from instrumentation basics to recommended controls and data analysis.