We have found 90-95% of commercially available mAbs work fine on 4% PFA fixed cells that are permeabilized with saponin. I interpret this to mean that most mAbs recognize epitopes that are not denatured by 4% PFA. Although I do not have as extensive experience with the various whole blood lyse-fix reagents out there, I have noted several mAbs that work fine with 4% PFA, but do not with several of the commercial lyse-fix reagents that I have tried. My interpretation is that these fix-lyse reagents caused more denaturation, perhaps due to the use of a different aldehyde fixative, different pH, etc. My experience with the commercial fix-lyse reagents is by no means comprehensive. Bottom line: just use your favorite anti-HLA-DR. You have a 90-95% chance of getting nice staining with 4% PFA/saponin, maybe a little less with fix-lyse reagents. Protocol: Current Protocols in Immunology 6.24. > ---------- > From: THIERRY FUMEAUX > Sent: Tuesday, March 6, 2001 3:31 > To: Cytometry Mailing List > Subject: Intracellular HLA-DR FACS analysis > > > Does anyone have any ideas about intracellular staining and FACS analysis > for > HLA-DR in Human monocytes ? > > I would be interested in knowing what kind of antibodies to use (if any is > suitable)and what protocole > > Thank YOU > > ------------------------------------------------------- > Ce message a volontairement ete envoye sans accent pour > permettre une compatibilite avec toutes les plateformes > ------------------------------------------------------- > Dr Thierry Fumeaux > Chef de Clinique adjoint > Soins Intensifs de Medecine > Hopital Cantonal Univesitaire > Rue Micheli-du-Crest 24 > CH - 1211 GENEVE > 022 / 372 33 11 > 022 / 372 56 86 > > > ____________________________________________________________________ > Get free email and a permanent address at http://www.netaddress.com/?N=1 >
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