CORRELATION BETWEEN MONOCLONAL-ANTIBODY REACTIVITY AND EXPRESSION OF CD4 AND CD8-ALPHA GENES IN THE HORSE

Equine peripheral blood lymphocytes (PBL) were enriched by positive selection using panning with a mixture of monoclonal antibodies against putative equine CD4 (Equine Leucocyte Antigen Workshop antibodies WS 1 and WS 72), or CD8 molecules (Workshop antibodies WS 12, WS 49, and WS 74). RNA was extracted from CD4 enriched cells (99% purity), from CD8 enriched cells (69% purity), from peripheral blood lymphocytes, and from neonatal equine thymus. RNA extracted from equine granulocytes and from equine kidney served as negative control. The RNA was electrophoresed in agarose and transferred to nylon membranes. Northern blots were hybridized with human and mouse cDNA probes for CD4 and CD8 alpha. The human CD4 probe detected a 2.9 kb RNA transcript in equine PBL, CD4 enriched lymphocytes, and thymocytes. The human CD8 alpha probe detected a 2.0 kb transcript in RNA from equine CD8 alpha enriched lymphocytes and thymocytes, but not from PBL or CD4 enriched lymphocytes. Mouse cDNA probes for CD4 and CD8 did not react with equine RNA. Results of hybridizations using the human probes support the assignment of the CD4 and CD8 specificities to the antibodies listed above. The results also suggest that the equine CD4 and CD8 alpha genes are more closely related to the human than to the murine counterparts.