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>Raymond, CHANG asked about fluo-3 and calcium measurements.
We try to useFluo-3. In our studies, the cells were incubated in a temp- and
pH controlled chamber and we hope to measure the Ca2+ up to 90 mins, i.e. to
take the cells into the flow every 5 minutes for measurement. We wonder
the dye is suitable for such long time incubation at 37C. Is there anyone
who has such experience?
I have used Fluo-3 to measure Ca2+ in thymocytes over 80 minutes. This method
involved pre-incubating the cells for 10 mins at 2e7 cell/mL with dye at
6ug/mL of cells at 37oC. The sample was then diluted to 1e6 cell/mL and run
collecting data at 5 sec intervals. It worked very well. Larger cells may
need a higher concentration of fluo-3. I used thapsigargin at 10nM or 100nM
as a positive control each day to make sure the set up was still OK. It should
yield a large increase in intracellular calcium within a few minutes.
Jo Beaver
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CD-ROM Vol 3 was produced by Monica M. Shively and other staff at the
Purdue University Cytometry Laboratories and distributed free of charge
as an educational service to the cytometry community.
If you have any comments please direct them to
Dr. J. Paul Robinson, Professor & Director,
PUCL, Purdue University, West Lafayette, IN 47907.
Phone: (765)-494-0757;
FAX(765) 494-0517;
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