A direct immunoradiometric assay for human plasma prorenin: concentrations in cycling women and in women taking oral contraceptives.
Two synthetic penta deca peptides corresponding to the N-terminal portion (amino acid sequence 1-15) and the C-terminal portion (sequence 32-46) of the pro moiety of the human prorenin (PR) molecule were coupled to BSA and used as antigens to generate antibodies against PR. In a RIA system using the 125I-labeled peptide as tracer, it could be shown that antibodies against peptide 32-46 bound the peptide and the native PR in the follicular fluid (FF) to a similar degree, whereas antibodies generated against peptide 1-15 did not specifically recognize native PR. The specificity of the PR-(32-46) antibodies for PR was demonstrated by comparative measurements of PR by RIA and by an indirect procedure (involving trypsin treatment of PR) in different individual FF samples, plasma samples, and fractions of FF obtained by gel filtration or immunoaffinity chromatography. Normal plasma PR levels could not be measured by RIA, since they were below the detection limit of the assay (0.5 micrograms PR/L approximately 10 pmol/L). For measurement of the low PR levels in plasma, a sensitive direct immunoradiometric assay with a detection limit of 0.1 fmol PR/tube was developed. It was based on the combined action of a commercially available solid phase renin antibody and the affinity-purified and 125I-labeled PR-(32-46) antibody. The measurement of 35 individual plasma samples with different PR concentrations showed an excellent correlation (r = 0.99) between the new direct and the conventional indirect assays. The direct assay of PR concentrations in plasma of healthy women during the length of a menstrual cycle resulted in a biphasic pattern of PR concentrations, with peak levels (approximately 3-fold increase) at the time of the LH surge. The intake of monophasic and triphasic contraceptives caused a suppression of normal PR concentrations (96.9 +/- 34 ng/L; early follicular phase; n = 12) by 39% and 25%, respectively, which was also observed in the pill-free phase of the artificial cycle.
Schumacher
M
M
Institute for Hormone and Fertility Research, Hamburg, Germany.
Nanninga
A
A
Delfs
T
T
Mukhopadhyay
A K
AK
Leidenberger
F A
FA
D000328:Adult; D002846:Chromatography, Affinity; D003276:Contraceptives, Oral / Q000494:pharmacology*; D004792:Enzyme Precursors / Q000097:blood*; D005260:Female; D005498:Follicular Phase; D006801:Humans; D007106:Immune Sera; D015592:Immunoradiometric Assay / Q000379:methods*; D008597:Menstrual Cycle; D009994:Osmolar Concentration; D011863:Radioimmunoassay; D012083:Renin / Q000097:blood*
D003276:Contraceptives, Oral; D004792:Enzyme Precursors; D007106:Immune Sera; D012083:Renin