Chloromethyl-X-Rosamine is an aldehyde-fixable potential-sensitive fluorochrome for the detection of early apoptosis

Cytometry
A MachoG Kroemer

Abstract

Early apoptosis is invariably accompanied by a disruption of inner mitochondrial transmembrane potential (delta psi m). Cationic lipophilic fluorochromes, such as 3,3' dihexyloxacarbocyanine iodide (DiOC6(3)), rhodamine 123, or 5,5', 6,6'-tetrachloro-1,1', 3,3'-tetraethylbenzimidazolcarbocyanine iodide (JC-1), can be used to measure such an apoptotic delta psi m dissipation. However, these dyes are afflicted by the handicap that cytofluorometric analyses must be performed ad hoc on nonfixed, metabolically active cells. Here, we show that chloromethyl-X-rosamine (CMXRos) is a viable alternative to other delta psi m-sensitive probes, and that it allows for formaldehyde fixation of cells before analysis. Using this fluorochrome, we developed a three-color staining technique in which two fluorochromes (fluorescein isothiocyanate and phycoerythrin) coupled to antibodies are employed to determine expression of cell-surface antigens, and CMXRos is used to measure delta psi m. In addition, we developed an approach to assess simultaneously delta psi m and expression of intracellular antigens. Thus, the expression of Bc1-2, a mitochondrial outer-membrane protein, can be determined after staining with CMXRos, fixation, and cell permeabili...Continue Reading

Citations

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