Development of a homologous radioimmunoassay for rat relaxin.

Highly purified rat relaxin has been radioiodinated to specific activities of approximately 100 micro Ci/microgram with the Bolton and Hunter reagent [N-succinimidyl 3-(4-hydroxy-5-[125I]iodophenyl) propionate]. A rabbit antirat relaxin serum, applicable in a final dilution of 1:100,000, was developed in a rabbit using unconjugated highly purified rat relaxin. A specific and precise double antibody RIA for rat relaxin sufficiently sensitive to routinely measure from 32--2000 pg rat relaxin was developed. Using this RIA, relaxin immunoactivity levels in extracts of pregnant rat ovaries were found to rise from 0.8 microgram/geq ovarian fresh tissue on day 8 of pregnancy to 723 microgram/geq ovarian fresh tissue on day 20 of pregnancy and then to drop precipitously to 6 microgram/geq ovarian fresh tissue on day 1 of lactation. Consistent with the occurrence and relative levels of relaxin in the ovarian extracts, levels of relaxin in pregnant rat serum were less than 2 ng/ml on day 10 of pregnancy, approximately 150 ng/ml on days 20 and 22 of pregnancy, and 12 ng/ml on day 2 of lactation.

Radioimmunoassay of relaxin throughout pregnancy and during parturition in the rat.

Relaxin immunoactivaity concentrations in the peripheral sera been measured during pregnancy and parturition in rats with a homologous rat relaxin RIA. Relaxin was not detected until day 10 of pregnancy (day 10). Relaxin levels increased markedly over the next 4 days and generally ranged between 50-100 ng/ml from days 14-20. A prepartum surge in relaxin levels, which appeared to be associated with the photoperiod, occurred on day 21 in unanesthetized rats bled via indwelling jugular cannulas. Mean relaxin levels in these animals increased sharply from less than 80 ng/ml on day 20 to approximately 140 ng/ml at 1400 h on day 21 and declined to less than 60 ng/ml during the 12-24 h preceding parturition. The prepartum surge in relaxin immunoactivity may be associated with luteolysis, since there was an accelerated decline in progesterone concentrations concurrent with the prepartum relaxin surg. Rats also experienced a surge in relaxin concentrations during parturition. The mean maximal level of relaxin during parturition was approximately 180 ng/ml. The rate of decline of rat relaxin immunoactivity levels after bilateral ovariectomy on day 21 demonstrated characteristics of a multiexponential curve. The mean clearance t 1/2 from 0-30 min was 22 +/- 3 min (+/- SEM), and the mean t 1/2 from 30-180 min was 56 +/- 7 min (+/0 SEM).