Fig. 7

Functional differences in spermatozoa immobilization and aggregation by SPP120 (a) Seven SPP120 constructs were developed to investigate the function of each domain. (b) The effect of recombinant SPP120 on sperm motility was examined after the addition of expressed SPP (final concentration = 10 μg/ml). The percentage of mobile spermatozoa before and 30 s after the addition of SPP120 was calculated based on video recordings and statistically analyzed using Tukey’s HSD test (N = 5). (c) In the aggregation assay, spermatozoa were first suspended in 30 μl of a Ca²⁺-free activation solution containing expressed SPP120 (30 μg/ml); 3 μl of 100 mM CaCl₂ were then added to observe sperm aggregation (N = 5). Arrowheads show the aggregated-sperm. (d) In the cross-linking assay, 10 μg of each protein were applied in SDS-PAGE after cross-linking with or without 100 μM DSS, and detected using anti-SPP120 antibody