ANTI-BULL SPERM MONOCLONAL-ANTIBODIES .2. BINDING CHANGES DURING CAPACITATION AND INFLUENCE ON SPERM-ZONA INTERACTIONS IN-VITRO

Anti-bull sperm monoclonal antibodies (mAbs), generated against intra- acrosomal and surface antigens, were evaluated for their functional si gnificance. In experiment I, the influence of mAbs on the bovine sperm -oocyte interaction in vitro was tested on a total of 493 oocytes in e ither three or four replicate trials. Although the number of sperm bou nd per zona increased significantly over untreated control samples (23 .6 +/- 5.6 vs. 10.0 + 2.4, mean +/- standard error [SE]; P < 0.001) in the presence of one surface-reacting mAb, other mAbs had no effect. E xperiment II was designed to determine if the mAbs would detect capaci tation-related changes of bull sperm in vitro. Bull sperm were incubat ed in capacitation medium (supplemented Tyrode's medium [TALP] plus 10 mu g/ml heparin) for up to 4.5 hours. At 0 and at 4 hours, mAbs in hy bridoma culture supernatant were incubated for 30 minutes with sperm, labeled with fluorescein isothiocyanate (FITC)-conjugated secondary an tibody, and processed for indirect immunofluorescence assay. Four mAbs specific to intra-acrosomal antigens exhibited a time-dependent incre ase (P < 0.05) in binding to bull sperm incubated under capacitation c onditions. In contrast, the binding of the mAbs specific to surface an tigens significantly decreased (P < 0.05) after 4 hours in the presenc e of heparin. Sperm viability did not change during the 4-hour period. In experiment III, mAbs specific to intraacrosomal antigens were eval uated to assess bull sperm acrosomal status following lysophosphatidyl choline-induced acrosome reaction. A significant decrease (P < 0.01) i n mAb binding following the induced acrosome reaction was observed wit h all the mAbs; it was highly correlated (r greater than or equal to 0 .85; P < 0.01) with Pisum sativum agglutinin binding and Giemsa staini ng. The results suggest that some of the mAbs are potential biological markers for bull sperm surface changes associated with capacitation a nd the acrosome reaction in vitro.