Fig. 2. Increase in phospho-PKC substrate following motility initiation of intact sea urchin spermatozoa. The phosphorylation level, as detected by immunoblotting with an anti-phospho-PKC substrate antibody, of several proteins was markedly increased when immotile (i) L. pictus spermatozoa were diluted into ASW to initiate their motility (m) (). The increase in phosphorylation of four of these proteins (arrowheads at 200, 100, 65 and 28 kDa) was observed in all the experiments. The specificity of the antibody for these proteins is demonstrated by the disappearance of bands when the antibody was pre-incubated with its antigenic phospho-peptide (2261) but not after pre-incubation with a mixture of phospho-amino acids (phos-mix) or a different phospho-peptide (9621). A blot representative of two other blots is presented.