Radioimmunoassay of vasopressin during pregnancy. Use and removal of cystylaminopeptidase inhibitors.
This paper describes a radioimmunoassay (RIA) for arginine-vasopressin in which o-phenanthroline effectively inhibits cystyl-amino-peptidase activity in whole blood and plasma from pregnant women but in which o-phenanthroline is removed during the extraction of vasopressin from plasma to prevent disturbance of the RIA. Cystyl-amino-peptidase causes immediate degradation of vasopressin unless cystyl-amino-peptidase enzyme inhibitors such as o-phenanthroline are applied. However, o-phenanthroline interferes with RIA. We report an extraction procedure over octadecasyl silica-packed Sep-Pak C18 columns, by which cystyl-amino-peptidase as well as most of the cystyl-amino-peptidase inhibitor is removed from plasma with chloroform. The average o-phenanthroline concentration (0.25 mmol/l) found in the assay medium after extraction appeared not to interfere with the RIA. Polyacrylamide gel isoelectric focusing of extracts of platelet-rich and platelet-poor plasma from pregnant women revealed a single vasopressin immunoreactive peak in the RIA. Recovery and between-assay coefficients of variation of 3.2 ng/l vasopressin from pregnancy whole blood were comparable with non-pregnant controls (57%/8% and 59%/13%, respectively). Results with this assay compare well with those of another assay using inhibitors in pregnant subjects and with results in non-pregnant subjects.
van der Post
J A
JA
Department of Obstetrics and Gynecology, Academic Medical Center, University of Amsterdam, The Netherlands.
van Heerikhuize
J J
JJ
Boer
K
K
van Boxtel
C E
CE
Swaab
D F
DF
D001127:Arginine Vasopressin / Q000097:blood* / Q000302:isolation & purification; D010122:Cystinyl Aminopeptidase / Q000037:antagonists & inhibitors / Q000097:blood; D005260:Female; D006801:Humans; D007202:Indicators and Reagents; D007525:Isoelectric Focusing; D010618:Phenanthrolines / Q000494:pharmacology; D011247:Pregnancy / Q000097:blood*; D011863:Radioimmunoassay / Q000379:methods; D012016:Reference Values; D012680:Sensitivity and Specificity
D007202:Indicators and Reagents; D010618:Phenanthrolines; D001127:Arginine Vasopressin; D010122:Cystinyl Aminopeptidase; C025205:1,10-phenanthroline