Flow cyt. & in situ hybridized WBCs

From: David Matthes (dmatthes@email.sjsu.edu)
Date: Mon Mar 26 2001 - 20:49:25 EST


Based on EST analysis I have found that there are 24 murine homologs of the genes I
study.  No antibodies are currently available for most of them yet I know that some are
expressed by cells of the spleen, thymus or lymph node.  I would like to assess quickly
which leukocytes each of these genes are expressed in, and possibly get a rough idea of
how high that expresssion is.  It seemed like in situ hybridization to WBCs followed by
(preceded by?) antibody labeling and then flow cytometry might be one way to pursue this.

Though I suspect there is a technical reason why reports of cells being analyzed in
this way (by signal strength of fluorescent riboprobe- labeled cells) are not common
-- perhaps because of weak signal due to the diffusiveness of cytoplasmic mRNA? --
I thought there might be someone more expert in flow cytometric analysis who had
experience with this and could steer me to an article or protocol, or tell me what
the heretofore unresolved technical difficulties are.  Anyone?

David



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