Hi everybody, I am rather new to the field of flow cytometry and I failed already at finding two clearly distinguishable dyes for my experiments. I am trying to measure gap junctional coupling by loading one population of cells with a freely diffusable dye, which is calcein, AM (Molecular Probes). It is excited at 488 nm and emits light at about 520 nm. The problem now is to find a dye (or any other label which is traceable in a flow cytometer) which is excited at about the same wavelength (in our FACS Analyzer there is only one excitation wavelength) AND which labels a population of cells permanently and does not diffuse via gap junctions. By mixing a population of doubly stained cells as described above with a population of unstained cells I wanted to measure the amount of cells which receive calcein from the pre-loaded cells and show thus only one fluorescence. We already tried DiA and DiI (Molecular Probes) which were not suitable for the task. I would be glad for any advice I can get. Thanks a lot, Anja @@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@ Anja Mindermann Abt. Biophysik, Biologisches Institut, Universitaet Stuttgart, Pfaffenwaldring 57, 70550 Stuttgart Tel.: (+49)711/685-5028 oder -7481 E-Mail: ami@tebio1.biologie.uni-stuttgart.de anja.mindermann@po.uni-stuttgart.de @@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@@
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