Quantification of intact plasma AGT consisting of oxidized and reduced conformations using a modified ELISA.

Quantification of intact plasma AGT consisting of oxidized and reduced conformations using a modified ELISA. Am J Physiol Renal Physiol. 2016 Aug 10;:ajprenal.00320.2016 Authors: Satou R, Kobori H, Katsurada A, Miyata K, Navar LG Abstract The pleiotropic actions of the renin-angiotensin system (RAS) depend on the availability of angiotensinogen (AGT) which generates angiotensin I (Ang I) when cleaved by renin. Thus, quantification of the intact AGT (iAGT) concentrations is important to evaluate the actual renin substrate available. The iAGT conformation exists as oxidized AGT (oxi-AGT) and reduced AGT (red-AGT) in a disulfide bond, and oxi-AGT has a higher affinity for renin, which may exacerbate RAS associated diseases. Accordingly, we determined iAGT, oxi-AGT and red-AGT levels in plasma from rats and mice. Blood samples were obtained by cardiac puncture, then immediately mixed with an inhibitor solution containing a renin inhibitor. Total AGT (tAGT) levels were measured by tAGT ELISA which detects both cleaved and iAGT. iAGT levels were determined by iAGT ELISA which was found to only detect red-AGT. Thus, it was necessary to treat samples with dithiothreitol, a reducing agent, to quantify total iAGT concentration. tAGT levels in rat and mouse plasma were 1839±139 and 1082±77 ng/ml, respectively. iAGT levels were 53% of tAGT in rat plasma but only 22% in mouse plasma, probaby reflecting the greater plasma renin activity in mice....
Source: Am J Physiol Renal P... - Category: Urology & Nephrology Authors: Tags: Am J Physiol Renal Physiol Source Type: research