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> Can anyone recommend protocols and reagents for enumeration of human CD34+
> cells in cord blood, also in cytokine/factor stimulated cultured leucocytes from
> different sources. I am using a BD FACS Sort with a Mac (CellQuest software),
It depends on exactly what questions you're trying to answer. For
strictly counting CD34+ cells, the combination of PE-labeled anti CD34
(BD's HPCA-2 is probably the best product) and CD45-FITC is certainly a
good combination, remembering that blast cells have mid-range CD45
expression. Light scattering parameters are also very helpful, as CD34+
cells have a highly reproducible light scatter profile. The biggest
mistake people make in enumerating CD34 cells is counting cells with
middling expression- like second log by PE. This is usually
non-specific, often due to monocytes, and is easily resoved with the CD45
double label.
I am also new to the list, btw. Glad to have found it.
Steve Gore
Johns Hopkins Oncology Center
Balto, MD
410-955-8781
sdgore@welchlink.welch.jhu.edu
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