Amniotic fluid (AF) from 25 term pregnancies was analyzed for cyclo(His-Pro)-like immunoreactivity (CHP-LI). CHP-LI was detected in all AF samples and was indistinguishable from synthetic CHP by immunoidentity, by gel chromatography on Sephadex G-25, and by high pressure liquid chromatography. The mean concentration of CHP-LI in AF was 13,622 +/- 1288 pg/ml (+/- SE) and concentrations were not altered by maternal labor. Plasma concentrations of CHP-LI were similar in 4 pregnant and 4 control subjects [2260 +/- 432 pg/ml vs. 2162 +/- 419 pg/ml (+/- SE), respectively]. We conclude that 1) CHP-LI is readily detected in AF from term pregnancies and is indistinguishable from synthetic CHP, and 2) concentrations of CHP-LI in human AF are significantly higher than concentrations of maternal plasma CHP-LI, suggesting CHP AF originates by mechanisms other than diffusion from maternal plasma.

A radioimmunoassay for cyclo(His-Pro) (CHP) in unextracted human plasma that can detect 40 pg has been developed. Elution profile of CHP-like immunoreactivity (CHP-LI) corresponded precisely to those of [3H] cyclo(His-Pro) by both high pressure liquid chromatography and Sephadex G-25 column chromatography. In addition plasma CHP-LI exhibited close immunoidentity with authentic CHP. Charcoal treatment of plasma containing [3H] cyclo(His-Pro) resulted in loss of both CHP-LI and [3H] cyclo(His-Pro) activity. Plasma frozen at draw and assayed at 0,6 and 24 hours displayed no change in CHP-LI while an aliquot from the same sample maintained at 4°C and assayed at the same time intervals showed a 50% rise (0 hrs-856 ± 47 pg/ml; 24 hrs-1288 ± 85 pg/ml) (±SE) over 24 hours. In addition, plasma drawn from 14 volunteers and immediately frozen until assay yielded a mean CHP value of 829 ± 64 pg/ml (±SE) while that of 14 volunteers that was maintained at 25°C for 90 minutes was significantly higher at 1085 ± 34 pg/ml (±SE) (p < 0.03). Taken together, these data suggest that CHP can be easily measured in a direct RIA of human plasma, however, failure to maintain the sample frozen from the time of draw until assay may yield spuriously elevated values.