Radioimmunoassay of plasma renin activity

Plasma renin activity is quantitated by measuring the rate of angiotensin generation during incubation of plasma renin with endogenous renin substrate. The angiotensin is quantitated by radioimmunoassay. Our studies indicate that the incubation step is best carried out in undiluted plasma at the pH optimum for renin (pH 5.7) in the presence of EDTA, neomycin, and DFP or PMSF. By using these conditions, incubation of low-renin samples can be prolonged for up to 18 hr, because angiotensinases and converting enzyme are completely inhibited. Accuracy is enhanced by prolongation of the incubation time, which results in more angiotensin and eliminates the need for blank subtraction. Incubation at pH 7.4 is disadvantageous because of lower rates of generation of angiotensin 1, because of the inability to maintain pH constant without addition of strong buffer, and because the incubation step cannot be prolonged beyond 3 hr. Dilution of plasma is undesirable because it results in a slower reaction rate due to dilution of both enzyme and substrate, and it is not possible to correct for the effect of substrate dilution. The radioimmunoassay of angiotensin I presents few unusual problems. The volume of plasma assayed should be 20 muI or less. If blank subtraction is used, antibodies should be screened to determine the extent to which they bind nonspecific substances in plasma, and then to ascertain whether the blank is entirely additive when angiotensin is added to it. Assay sensitivity is an important issue, since approximately 30 percent of patients with essential hypertension have subnormal plasma renin activity. In a study of three different commercial kits we found that many low-renin samples were undetectable and major fractions could not be discriminated with precision or consistency from normal renin samples. However, the incubation conditions can be easily modified, so that an 18-hr incubation can be utilized and then low-renin samples can be detected with the same degree of accuracy as those with normal plasma renin activity.