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It is widely recognized that fluorescent labeling of cells is an effective means to determine total cell numbers or how many viable cells exist in a sample. Flow cytometry combined with fluorescent staining is a powerful tool to analyze heterogeneous cell populations. Fluorescein diacetate (FDA) and its derivatives are non-fluorescent molecules that diffuse into cells and are hydrolyzed by intracellular non-specific esterases to give fluorescent products. The fluorescent products can be accumulated only in those cells that have intact cell membranes; therefore, dead cells with leaky membranes are not stained. The precise kinetics of membrane transport and intracellular hydrolysis of FDA and its analogs (such as CDCFDA) are related to cellular functions, thus FDA labeling can be used for monitoring cells by flow cytometry or fluorescence microscopy. The fluorescence intensity of labeled cells by FDA dyes varies considerably among cell lines and strains, probably because of differences in intracellular esterase activity. CFSE is an amine-reactive FDA derivative that is widely used for monitoring cell proliferation with a flow cytometer.
CalculatorsCommon stock solution preparation
Table 1. Volume of DMSO needed to reconstitute specific mass of CFSE [5-(and 6)-Carboxyfluorescein diacetate, succinimidyl ester] *CAS 150347-59-4* to given concentration. Note that volume is only for preparing stock solution. Refer to sample experimental protocol for appropriate experimental/physiological buffers.
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