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Ray:
I have had luck with in detecting PKC isoenzymes via flow
cytometry in NK cells and other lymphocytes fixed by the
following procedure:
1. Cells in microplate wells (96 V-well plate, 1-2x106 cells per well)
are washed twice by centrifugation (400 g, 5 min, 4C) in phosphate
-buffered saline (PBS), pH 7.2.
2. Resuspend cells in 200 microL PBS containing 4% (w/v)
paraformaldehyde (freshly prepared) and incubate at 37C for 15 min.
3. Wash cells in 200 microL cold (4C) 0.02% (v/v) Triton X-100 in PBS.
4. Wash next in 200 microL PBS containing 1% (v/v) Tween-20 and 1%
bovine serum albumin, pH 7.8.
5. Suspend cells in PBS containing 0.2% bovine serum albumin for
immunofluorescent staining.
I have not investigated use of gluteraldehyde.
Good luck
*************************************************************
Bruce Edwards
Scientist
The Lovelace Institutes
Albuquerque, NM 87108
Email: Bruce@lucy.tli.org
Phone: 505-262-7155
Fax: 505-262-7043
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