Greetings to all,
I initially posted a response privately to this question, but I would
like to ask a general question of those much more knowledgeable than I.
Until recently I had assumed that pressure, sheath fluid, and collection
medium were the main causes of cell death after sorting. However, recently
we had a problem with stray voltage on a FACStarPlus (shocks after just
touching the cytometer, plates charging by themselves, etc.) that was
coincident with lousy viability after sorting. In frustration, I performed
the following: 1) I collected the waste stream (pressure, no light, no
electrical field)--cells were 97% viable, 2) I collected the waste stream
(pressure, laser engaged, plates on)--cells were 96% viable. 3) I sorted
cells--10% viable! (All of these tasks were performed in less than a 5
minute period). We had zapped the cells. Is it possible that the
electrical charge behaves somewhat like electroporation which is really
hard on cells? The manufacturer was stumped, but a retooling of the
instrument solved the problem. I have never seen a discussion on this
aspect of sorting, but could this also contribute to loss of viability and
function? If so, can we adjust this?
Thanks,
Kathy
--------------------------------------------------------------
Kathy Schell
Supervisor, UWCCC Flow Cytometry Facility
600 Highland Ave. K4/535
Madison, WI 53792
Voice: 608-263-0313
e-mail: kschell@facstaff.wisc.edu
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