Motilin-antimotilin reaction reveals two antigenic determinants in motilin.

The reaction of motilin with four rabbit antimotilin sera raised by immunization with synthetic porcine motilin-bovine serum albumin conjugate was studied with respect to various binding parameters and specificity. All four antisera exhibited an extremely high degree of specificity and high affinity (K greater than 10(11) M-1) for porcine motilin. Studying the various synthetic motilin fragments. These antisera appeared to contain binding sites reacting strongly with the N-terminal sequence in which the first three amino acid residues are essential for high affinity binding. One of the antisera, R-3-6, appeared to contain approximately 20% of its binding sites with high affinity for C-terminus-containing fragments. These results suggest that motilin possesses two antigenic domains along its primary structure; one contains the N-terminal tripeptide and the other contains the C-terminal nanopeptide as essential parts. Thus, in addition to heterogeneity in affinity, a given antibody preparation may be heterogenous with respect to the specificity along the sequence of a peptide. Gel filtration studies of the methanol extracts of human and dog plasma indicated that an immunoreactive motilin-like material with a molecular size similar to natural porcine motilin was measured by our routine radioimmunoassay.