Fig. 8. NSF is present in isolated planar cortex but is not associated with CVs. (A) Replicate western blots probed for sea urchin NSF, calreticulin and two CV cargo proteins. Lane 1, IPC containing 1.5x10⁷ CVs; lane 2, an equivalent amount of cortex denuded of CVs; lanes 3-5, 1.5x10⁷ isolated CVs (three different preparations); lane 6, egg cytosol (5 µg total protein); lane 7, egg microsomes (10 µg total protein). (B) NSF western blot. Lane 1, egg cytosol (5 µg total protein); lanes 2-6, recombinant NSF-His₆ standard curve (0.48, 0.24, 0.12, 0.048, and 0.024 fmol as hexamer); lanes 7, 8, two preparations of 1.5x10⁷ isolated CVs. (C) Recovery in western blots of recombinant sea urchin NSF added to cortical vesicle material. Lane 1, 0.024 fmol NSF alone; lane 2, 0.048 fmol NSF; lane 3, CV proteins; lane 4, CV proteins plus 0.024 fmol NSF; lane 5, CV proteins plus 0.048 fmol NSF. Representative of two experiments.