back on mouse T cells calcium flux

DAL SECCO Valentina ICH valentina.dal_secco at humanitas.it
Tue Jan 22 05:43:29 EST 2008


Dear FLOWers, thank you very much for your suggestions!

Ok, after many attempts, I agree that Fura 2-AM gives a lot of trouble and
is not useful for studying calcium flux responses of mouse naïve T cells. I
have managed to improve the signal, I mean the sensitivity of the
fluorimeter, and get good results only with cells I used as positive
controls such as Jurkat cells.

It might look obvious but was not: as for the Facs, you must set the Voltage
as best for the type of cells you are using!

 

Anyway I moved to FACS with Fura Red and Fluo-4. I have a question about
this:

 

Do you think I could detect any calcium flux by challenging with a stimulus
mouse naïve T cells within which responding cells would be only 20% of the
whole population?

 

I Know it would be much better to first select this subset and get a
population in which all cells would respond; however, to do so I should make
a positive selection by using an antibody against specific Tg TCR just
before using them; therefore I expect some unspecific fluxing to occur or
even specific flux to be diminished. Am I correct?

 

Thanks a lot for any suggestions!!!

Valentina

 

 

-------------- next part --------------
HTML attachment scrubbed and removed


More information about the Cytometry mailing list