Re: Post Sort Quenching
BIGOS@Darwin.Stanford.EDU
Mon, 10 Feb 1997 08:22:28 +0000 (PDST)
Dave Parks and I spent some time looking at loss of
fluorescence of PE to try to figure out why low power
Fascans did better than high power jet-in-air sorters at
measuring PE stained cells. We determined the PE
saturation curve and found that the dye saturated
significatly at 100mW. Moreover, we observed that with
several hundred milliwatts of 488nm illumination, PE was
kicked into some unusal state from which it didn't
recover. Re-analysis of sorted cells minutes later had
lost up to 45% of their origional fluorescence. With
less laser illumination, this effect was minimized. For
FITC we did not notice much change - 5% at most. We did
not look at Cy5PE at that time.
>From our experience we would expect that with the low
laser power that you are using (15mW) the losses for
FITC and PE will be small.
-Marty Bigos
Stanford Shared FACS Facility