An alternative strategy for the radioimmunoassay of angiotensin peptides using amino-terminal-directed antisera: measurement of eight angiotensin peptides in human plasma


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Abstract

We describe here a method of measuring angiotensin peptides and their carboxy-truncated metabolites in human plasma using N-terminal-directed antisera. Antisera raised against N-acetylated angiotensin (Ang) II and N-acetylated Ang III analogues were used to develop two radioimmunoassays. Extracted plasma samples were acetylated prior to separation of cross-reacting angiotensin peptides by high-performance liquid chromatography (HPLC). Fractions were assayed with both antisera to obtain measurements for eight angiotensin peptides. Angiotensin levels measured in normal males were (fmol/ml plasma, means ± s.e.m., n=14): Ang-(1—7) 1.0 ± 0.2, Ang II 13.9 ±2.0, Ang-(1-9) 7lt;0.4, Ang I 19.5 ± 2.4, Ang-(2-7) <1.1, Ang III 2.9 ± 1.0, Ang-(2-9) <2.1, Ang-(2-10) 2.4 ± 0.8. Hypertensive patients receiving angiotensin converting enzyme (ACE) inhibitor therapy (n=8) had an increase in Angt to 187.3 ± 107.2fmol/ml (P=0.002), and a reduction in Angll to 4.8 ± 1.2fmol/ml (P<0.001). Furthermore, these patients showed a ninefold increase in Ang-(1—7) to 9.7 ± 4.3 fmol/ml (P<0.001), indicating a role for prolylendopeptidase in the metabolism of Ang I in vivo. These N-terminal assays have demonstrated that carboxy-truncated metabolites of Ang I and Angll make little contribution to plasma angiotensin peptides, except during ACE inhibitor therapy. Furthermore, these antisera allow the measurement of Ang I and Angll in the same radioimmunoassay of fractions from HPLC, providing a highly reliable estimate of the Angll: Ang I ratio.

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