FUNCTIONAL AND PHENOTYPIC CHARACTERIZATION OF MONOCLONAL-ANTIBODIES TO BOVINE L-SELECTIN

Objective-To characterize 2 bovine neutrophil monoclonal antibodies (M AB) as to effects on bovine neutrophil function and their binding anti gens on the cell surface of bovine neutrophils. Animals-16 healthy, la ctating Holstein cattle, 1 calf with leukocyte adhesion deficiency, an d 1 age-matched control calf, 2 healthy ewes, and 2 healthy human bein gs as neutrophil sources. Procedure-Neutrophil chemotactic and respira tory burst activities and calcium influx, and binding properties of th e 2 MAB were determined. Molecular mass of corresponding cell surface antigens also was determined, as was binding of human L-selectin MAB D REG56 to molecules recognized by MAB 11G10 and 2G8 on the surface of b ovine neutrophils. Results-MAB 11G10 and 2G8 inhibited chemotactic act ivity of bovine neutrophils, up-regulated amplitude of native chemilum inescence, and shortened the time to reach maximal chemiluminescence i nduced by serum-opsonized zymosan. Crosslinking both MAB with a second antibody induced rapid increase in intracellular free calcium concent ration. Binding density of MAB 11G10 and 2G8 to bovine neutrophils tre ated with trypsin was increased (P < 0.05), compared with that of untr eated neutrophils. Neutrophils treated with phosphatidylinositol-speci fic phospholipase C had decreased (P < 0.05) binding density of MAB 11 G10 and 2G8. Binding of the various MAB to neutrophils from calves wit h bovine leukocyte adhesion deficiency was lower (P < 0.05) than bindi ng to neutrophils from healthy calves. Expression of antigens recogniz ed by the aforementioned MAB on the surface of bovine neutrophils was decreased (P < 0.05) within 10 minutes. Conclusion-MAB 11G10 and 2G8 r ecognized L-selectin molecules on bovine neutrophil membrane. L-Select in (CD62L) is involved in low-affinity adhesion reactions between leuk ocytes and L-selectin ligand on postcapillary venular endothelial cell s.