Hello Israel, When is your colleague adding Brefeldin A (or Monensin) to the 3 day assay culture? It can be harmful to the cells to maintain these inhibitors in cell culture for more than 8-10 hours.Typically, it is better to add these inhibitors for the last 4-5 hours of the assay. This may not be the problem, but worth checking. thanks, Achal Pashine Chiron Corporation On Fri, 18 Feb 2005 10:29:41 -0600, Israel Collier <collieri@slu.edu> wrote: > Hey FLOWers, > > My colleague is performing a 3-day assay (Intracellular Cytokine) on the > FACScan. She is unable to maintain cell viability. After Day 1 of the > protocol, 70% of the cells are dying! What is she doing wrong--if > anything? Could it be the way the lymphocytes are initially processed? > I have attached the lab protocol. Thanks in advance. > > Fellow FLOWer, > Israel > > > This attachment - 'Procedure for Setting Compensation for Multi color analysis.doc' - > 23.55 KBytes - can be viewed at > http://www.cyto.purdue.edu/MD-parts/9bc9f67e5e45d84fe4d0e73e24a98e0349fb19ac.doc > > > This attachment - 'Protocol for staining intracellular cytokine_GP100TRP2.doc' - 38.40 > KBytes - can be viewed at > http://www.cyto.purdue.edu/MD-parts/e5b210b725d2b4539d352c9d4e2c2aa10a20211a.doc > > > This attachment - 'Protocol for staining intracellular cytokine_GP100TRP2-new.doc' - > 51.71 KBytes - can be viewed at > http://www.cyto.purdue.edu/MD-parts/2398de2ba87605d61beef019a0be35faf57cb7f6.doc > >Received on Wed Feb 23 12:38:00 2005
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