Figure 2: ( a ) Behavior of CRISP3 during human sperm capacitation and the acrosome reaction. Total protein extracts of fresh (F), capacitated (C) and ionophore-treated (C + I) sperm were analyzed by SDS-PAGE and Western blots with an anti-hCRISP3 antibody (α-CRISP3). ( b ) Localization of CRISP3 in human spermatozoa. Representative phase contrast (upper panels) and fluorescence (lower panels) microphotographs of human capacitated (C) and ionophore-treated (C + I) spermatozoa subjected to immunofluorescence labeling with α-CRISP3 or normal IgG as primary antibodies. ( c ) Relevance of CRISP3 for human sperm fusion ability. Capacitated spermatozoa were incubated for 15 min in medium containing either normal IgG, α-CRISP3-, α-CRISP1- or α-CRISP2-antibodies, then co-incubated with ZP-free hamster oocytes for 2.5 h and the percentage of penetrated oocytes was determined. Results represent the mean ± s.e.m. of at least three independent experiments, *P < 0.05 versus IgG.