Fig. 7. Ca²⁺ release activity in PLC-immunodepleted soluble sperm extracts. (A) Immunoblot analysis of PLC protein in hamster sperm extract supernatants after incubation with control IgG or anti-PLC antibody (S- and S+, respectively) and the corresponding precipitated proteins bound to control IgG beads or anti-PLC beads (P- and P+, respectively). (B) Ca²⁺ release activity of antibody-treated sperm supernatants, S– and S+, assayed fluorometrically in sea urchin egg homogenates. Scale bars indicate time (seconds) and relative fluorescence units (RFU). Arrows indicate time of addition (C) Ca²⁺ changes in fura-red loaded mouse eggs after microinjection with sperm extract that was either untreated (top trace), control IgG-treated (second trace, n=13) or anti-PLC antibody-treated (bottom two traces, n=13). In 6/13 cases (third trace), the anti-PLC antibody-treated sperm extract showed an injection artifact-related single Ca²⁺ spike; in other cases there was no Ca²⁺ change (fourth trace).