Thomas, Your osmotic swelling theory sounds very plausible to me. The Fl1 shift can also be explained by the mixing of the PBS sheath and PBS the cells are suspended in since Fitc is very PH sensitive. The PBS the cells are suspened in may not be buffered strongly enough. See the Molecular Probes catelog for more info. on how Fitc intensity changes with PH. Good luck! Glenn MIT >Hi there! > >Hope you don't mind a novice's question. >Using a FACSort in a simple single-staining experiment of human myoblasts I >observed that the cell population in the FSC/FL3 blot would "move" during >the course of a measurement towards higher FSC values only to stabilze after >about 10 secs. of measurement (ca. 100 events per s). In parallel the FL1 >(cells stained w. FITC-labelled sec. abx) histogramm would shift to higher >values also. >Could anyone explain why this happens? I was suggested osmotic swelling of >cells due to an imbalance between the PBS the cells are suspended in and the >PBS in the source tank. But this would not explein the FL1-shift, would it? > >Thanks in advance, > >Thomas
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