Defining bioassay conditions to evaluate sperm/zona interaction: Inhibition of zona binding mediated by solubilized human zona pellucida
Objective: Our goal was to study the influence of solubilized human zonae pellucidae on zona binding potential and acrosome reaction. Materials and Methods: Zona pellucida (ZP) solutions were prepared by dissolving zona in acidic buffer, NaH2PO4 (pH 2.5), to obtain 0.1 and 0.5 zona pellucida/μl. Zona binding capacity was evaluated by the addition of oocytes (10-fold) to sperm/zona pellucida solution droplets. The number of sperm bound to each oocyte was recorded. Zona pellucida mediated acrosome activity was evaluated after 60 min of coincubation of sperm and 0.5 ZP/μl. Results: The mean (±SE) number of sperm bound for control, 0.1 ZP/μl, and 0.5 ZP/μl was 181.2 ± 12, 79.6 ± 5, and 38.8 ± 3, respectively. Zona pellucida-exposed sperm populations showed significant more acrosome-reacted sperm compared to control sperm, namely, 78 versus 32%, respectively (P = 0.001). Conclusions: The observed zona binding inhibition might be ascribed to zona receptor blocking on the sperm surface or the inability of acrosome-reacted sperm to bind to the zona pellucida.