Flow Cytometric Analysis of Cell Division by Dye Dilution

The technique described in this unit uses the intracellular fluorescent label carboxyfluorescein diacetate succinimidyl ester (CFSE) to tag proliferating cells. Covalently bound CFSE is divided equally between daughter cells, allowing discrimination of successive rounds of cell division. The technique is applicable to in vitro cell division as well as to in vivo division of adoptively transferred cells and can resolve up to eight successive generations. CFSE is long lived, permitting analysis for several months after transfer, and has the same spectral characteristics as fluorescein, so monoclonal antibodies conjugated to phycoerythrin or other compatible fluorochromes may be used to immunophenotype the dividing cells.