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Canine CD34: cloning of the cDNA and evaluation of an antiserum to
recombinant protein
PA McSweeney, KA Rouleau, R Storb, L Bolles, PM Wallace, M Beauchamp, L Krizanac-Bengez, P Moore, G Sale, B Sandmaier, T de Revel, FR Appelbaum and RA Nash
Clinical Research Division, Fred Hutchinson Cancer Research Center,
Seattle, WA 98104, USA.
Increasingly, enriched populations of hematopoietic progenitors are used in
experimental and clinical transplantation studies. The separation of
progenitors is based on the expression of CD34, a marker preferentially
expressed on progenitor cells. The dog model has been important for
preclinical transplant studies, because it has proven predictive for
outcomes in human hematopoietic stem cell transplantation. To identify and
isolate canine hematopoietic progenitors, we have cloned a cDNA encoding a
CD34 homologue from a canine myelomonocytic leukemia cell line, ML2. The
CD34 homologue cDNA predicts an amino acid sequence that is highly
conserved with human and murine CD34 in the cytoplasmic domain,
transmembrane domain, and C- terminal end of the extracellular domain, but
shows considerable divergence from these sequences at the amino-terminal
end of the protein. In Western blotting studies, canine CD34 homologue
(caCD34) appears to be a heavily and variably glycosylated protein with a
molecular weight of approximately 100 kD and shows some tissue-specific
differences in protein mass. To evaluate the expression of caCD34 protein,
the extracellular domain of caCD34 was expressed as an Ig fusion protein
and used as an immunogen to generate a rabbit polyclonal antiserum. The
antiserum reacted against the fusion protein, against vascular endothelium,
and with three leukemic cell lines. Approximately 1% of canine bone marrow
cells stained brightly with antibodies to caCD34 and this population was
25- to 50-fold enriched for colony- forming units-granulocyte-macrophage as
compared to unfractionated marrow mononuclear cells. These findings suggest
that the canine CD34 homologue is expressed on bone marrow progenitor cells
and, thus, that this molecule should be a valuable marker for identifying
and isolating canine hematopoietic progenitors for experimental
hematopoiesis and stem cell transplantation.
Volume 88,
Issue 6,
pp. 1992-2003,
09/15/1996
Copyright © 1996 by The American Society of Hematology
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