Rodent thymocyte suspension
Ralph Smialowicz 541-5776 (SMIALOWICZ@AM.HERL.EPA.GOV)
Wed, 9 Aug 1995 09:10:10 EST
I would appreciate any help with a problem that we have had in preparing
"clump free" thymocyte suspensions, from mice and rats, for flow
analysis. We routinely prepare thymocyte suspensions in RPMI followed by
centrifugation at 200 x g. Cell pellets are resuspended in PBS with 2% FBS,
0.1% sodium azide and 0.025% DNase, the latter is added to "prevent"
clumping. The cells are subsequently centrifuged and resuspended in PBS
with 2% FBS and 0.1% sodium azide for staining. If we have a clumping
problem, it usually occurs after the first wash. In general, the clumping
appears to be associated with cell suspensions with higher than usual cell
numbers. Any suggestions would be greatly appreciated. Thanks.
Ralph J. Smialowicz
US EPA
RTP,NC
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