Herstellung und Charakterisierung monoklonaler Antikörper gegen equine Leukozyten

Production and characterization of monoclonal antibodies specific
for equine leucocytes This work was initiated to produce new
monoclonal antibodies for the characterization of equine
leucocyte-subpopulations. For immunisation, leucocytes isolated
from the peripherial blood, lymph nodes and the intestinal
epithelium of horses, have been used. Out of six cell-fusions 78
clons were isolated. To characterize the mAbs, the attention was
focused on immunofluorescence analysis. Antibodies of the clones
4-13, 4-58, 1-19 and 6-39 have been purified by protein
G-sepharoses chromatography and, except mAb 4-58, biotinylated. MAb
4-13, mAb 4-58 and mAb 5-50 are binding specifically to equine
T-cells. Flow cytometer analyses demonstrated an increased
frequency following the stimulation with Concanavalin A. The
frequency of mAb 4-13- and mAb 4-58-positive cells matched the
total frequency of CD4- and CD8-positive cells. The number of
cells, marked by mAb 5-50, is slightly lower. Most likely these
three mAbs recognize three different, as yet unknown antigens. MAb
4-58 stimulated the in vitro proliferation of leucocytes. Moreover
mRNA for interleukin-4 and interferon gamma was detected in these
cells after the stimulation. By contrast the antibodies 1-19, 2-52,
4-36, 4-55 and 6-39 are B-cell-specific, since they were present on
cells that were Ig-positive. The monoclonal antibodies 6-5 and 6-17
stain all equine leucocytes. Possibly they are binding to an equine
CD45-molecule. The antibodies of clone 4-18 recognize an antigen on
granulocyctes. Preliminary data suggests that mab 4-39 is binding
to a lektin, that can be found mainly on B-Lymphocytes and
endothelial cells. No group of leucocytes and no antigen could be
assigned to the other antibodies. The antibodies described here
will be valuable tools to characterize different equine
leucocyte-subpopulations by flow cytometry.