Bcl-2 family proteins are intracellular antigens and thus their detection by indirect immunofluorescence and subsequent analysis by flow cytometry (FACS)
requires permeabilization of the cells using non-ionic detergents. Procedures are described here specifically for FACS analysis of BcI-2 but are readily adapted to detection of other Bcl-2 family members by adjustment of the choice and amount of primary antibody employed. The immunodetection of Bcl-2 or other Bcl-2 family proteins can be combined with traditional cell-surface marker analysis, as well as with DNA content analysis or TUNEL assays for detecting cells with fragmented DNA, in two- or three-colour FACS assays.
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