I found that ro/ra give different staining patterns dependent on manufacturer,
concentration, choice of fluorochrome and lysis method. I settled for FACS
lyse, CD45ro-PE from DAKO. I report
8+45ro-
8+45ro+
8++45ro-
8++45ro+
8++45ro++
I have not done that together with chemokines.
Regards
Gerhard
-----Original Message-----
From: bunny [SMTP:bunny@cotleur.com]
Sent: Tuesday, June 26, 2001 4:52 PM
To: Cytometry Mailing List
Subject: (human) CD8 analysis questions
friends-
I have a dilemma. I'm participating in a study analyzing
chemokine receptors in MS patients in clinical trials. The
combinations used are: CD4+/CD45RA+/CHEMOKINE and
CD8+/CD45RA+/CHEMOKINE.
Here is my question: for measuring the CD8/45RA portion, do
I set the markers on background (isotype or
unstained,whatever- that's not my current question) and
accept ALL CD8+ (DIM AND BRIGHT), or specifically CD8 bright+/CD45RA+.
For most of the chemokines, the results are the same (for
either /CD8+ bright/45RA+ or CD8+ dim/45RA+). But for one
marker the results are dramatically different.
{I do understand that the CD8 dims's usually contain NK
cells. I searched both cytometry archives and pubmed, and
found many valuable discussions of the roles of these
various subpopulations. For this study, adding more/or
changing CD markers is out of the question.}
So my main questions here are:
1)With the combinations described above, WHERE do people
normally set the marker for CD8+ cells;
and
2) do you then report the data as "%chemokine receptor
expression in activated CD8+45RA+ cells" ? or CD8+bright/CD45RA+
--
Bunny Cotleur
Cleveland Clinic Foundation
Neurosciences NC30
9500 Euclid Avenue
Cleveland, OH 44195
(216) 444-1164
cotleua@ccf.org
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