USE OF SYNTHETIC CANINE RELAXIN TO DEVELOP A RAPID HOMOLOGOUS RADIOIMMUNOASSAY

Canine relaxin (cRlx) was synthesized by a combination of solid-phase methods and sequential site-directed disulfide bond formation. Proof t hat the intended molecule had been synthesized was obtained by analyti cal HPLC of the intact and reduced molecule, by amino acid and sequenc e analysis, and by receptor binding and in vivo mouse interpubic ligam ent bioassays. Antisera to synthetic cRlx were raised in six male rabb its; these cross-reacted with relaxins of other species, but not with insulin, LH, FSH, hCG, or prolactin (PRL). Three of the antisera neutr alized relaxin-induced interpubic ligament formation in estrogen-prime d mice in vivo. A new homologous cRlx RIA was developed through the us e of rabbit antiserum 79888, synthetic cRlx for standards and I-125-la beled trace, and a goat anti-rabbit IgG-polyethylene glycol precipitan t. The new RIA can be completed in 26 h and has a sensitivity of 0.195 -0.39 ng cRlx/tube. Intra- and interassay coefficients of variation we re 3% and 12.5%. During pregnancy in bitches, serum cRlx rose to about 10 mu g/ml. Immunoactive cRlx was also detected in serum, colostrum, and milk of lactating bitches, but not in large volumes (100-300 mu l) of serum of pseudopregnant or estrous bitches. Immunoreactive cRlx wa s also found in seminal plasma, but not in serum, of male dogs. The ne w homologous cRlx RIA is simple, rapid, sensitive, and specific, and w ill be used in future studies of canine relaxin physiology.