Flow cytometry is the most widely used method for detecting and quantifying apoptosis in mammalian cells. The multiparametric nature of flow cytometry allows several apoptotic characteristics to be combined in a single sample, making it a powerful tool for analyzing the complex progression of apoptotic death. This chapter provides guidelines for combining single-apoptosis assays such as fluorogenic caspase substrates, annexin V binding, DNA dye exclusion, and covalent viability probes into informative multiparametric assays. This multiparametric approach to analyzing apoptosis provides much more information than single-parameter assays that provide only a percentage apoptotic result, given that multiple early, intermediate, and late apoptotic stages can be observed and quantified simultaneously. While much more informative than single-color assays, these multicolor methods can still be analyzed on relatively simple flow cytometers, making them accessible to many laboratories.
Keywords: 7-Aminoactinomycin D; Annexin V; Apoptosis; Caspase; Covalent viability probe; Flow cytometry; Fluorogenic caspase substrate; Hoechst dye; Pacific Blue; Propidium iodide.