APOPTOSIS IN INTERLEUKIN-3-DEPENDENT HEMATOPOIETIC-CELLS - QUANTIFICATION BY 2 FLOW CYTOMETRIC METHODS

The murine haemopoietic cell line, BAF3, undergoes apoptosis when the growth factor IL-3 is withdrawn. Two flow cytometric methods for quantifying the apoptotic cells are described. Cell sorting followed by DNA gel electrophoresis, and both light and electron microscopy have been used to identify the apoptotic cells. In the first method the cells are fixed in ethanol, stained with propidium iodide and a DNA histogram recorded. The apoptotic cells give a 'sub-G1' peak. In the second method unfixed cells are incubated with the bis-benzimidazole, Hoechst 33342. The apoptotic cells take up this dye more rapidly. In this latter method, the non-viable cells can also be enumerated by addition of propidium iodide. The value of the method has been demonstrated in a brief study of the effects of a panel of cytokines on growth and apoptosis.