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Cell concentration for flow cytometry - (May/15/2012 )

I am very new to flow cytometry and I am a bit confused with the issue on cell concentration for flow cytometry. Practically, do I have to count cell for every single sample and use the same cell concentration for each sample (e.g. 1 x 10^6 cells/100ul) for a specific amount of antibody (e.g. 5ul ab)?
If I use less cells then the set amount (e.g. 5 x 10^5 cells/100ul) with the same amount of antibody (e.g. 5ul ab), will there be variation on the results? (since the working dilution should be a saturating concentration, there should be no variation?)

-virginiakwan-

Hi,
Unfortunately yes , u have to count every single sample to get the correct concentration of cells
If u use the same ammount of antibody with different cell numbers u will have uncorrect results
Especialy if u use less cell number than the number u need
Usually, we take specific number of cells for every sample
For example (1x 10 ^5)
If the concentration of the sample is high u can dilute
But if the concentration is low u can centrifuge
I hope this helps

-Shadan-

also your ur Ab pairing with target cells ratio might be altered if you stil use same amount of Ab conc after lessen the cells conc

-mjon-