Flow cytometry is a powerful technique that can analyze properties of individual cells and measure millions of cells at a time. Single cell analysis typically includes measurements of surface and intracellular proteins, but protein modifications can also be detected by flow cytometry. Phosphoflow assays measure phosphorylated proteins and offers several advantages over traditional lysate-based phosphorylation detection assays.
- Single cell detection: Phosphoflow assays use fluorescently labeled antibodies that can bind to phosphorylated sites on proteins on the cell surface or within subcellular compartments. This means phosphorylation within individual cells can be measured
- Part of a panel: Like all flow cytometry assays, phosphoflow assays use a panel of fluorescently labeled antibodies to label different cell markers, thus allowing for identification of different cell subsets and analysis of phosphorylated proteins within these subsets. This type of assay can be used on different types of samples like whole blood and PBMCs and does not require additional cell enrichment steps to pull out rare cell populations
- Frozen in time: Protein phosphorylation is typically a transient modification, but specialized reagents have been developed for phosphoflow to fix proteins and preserve phosphorylation.
Phosphoflow analysis is valuable to understanding cell signaling events, and this can provide critical insights into basic cell functions and inform preclinical development of drugs and biologics. Even if phosphorylation levels are relatively low within individual cells, phosphoflow analysis across a cell population can detect phosphorylation events related to different cell signaling pathways. Consider adding phosphoflow to your next flow cytometry experiment.