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> Does anyone out there have a protocol for using sodium borohydride? I
> want to try to use it to reduce autofluorescence (Cytometry
> 8:235-239(1987)).
I'll be very interested in your results. I tried using thiosulfate and
sulfite for chemical reduction following the paper you cite (and some
others I think) but got less than 2-fold drop in autofluorescence of
EBV LCL (human B cells, JY line) even with concentrations which clearly
did quite a bit of damage to the cell structure. An idea given to me
by Janos Szollosi is to prebleach the cells with an intense 488 nm
laser beam (spread enough with a lens to cover a test tube), but I
haven't had the guts (retinal fortitude?) to try that. We tried 15
min or so in direct sunlight but that also did almost nothing. Probably
the best idea is to use a spare photomultiplier (if you have one) to
measure autofluorescence in a channel not being used for a fluorochrome.
This can be used to subtract autofluorescence from other channels quite
nicely as mentioned by someone to this list recently. There are a number
of references available on variants of this technique. Searching MEDLINE
for autofluroescence is a good way to get them (back 10 years or so).
/*- - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - - -
Eric Martz, Professor of Immunology emartz@microbio.umass.edu
Dept Microbiology Voice: 413-545-2325 FAX: 413-545-1578
Morrill IVN 203, Box 35720, Univ Massachusetts, Amherst MA 01003-5720
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