Biosynthesized [35S]methionine-labeled pro-opiomelanocortin peptides as novel recovery markers in radioimmunoassay of peptide hormones.

Abstract

Hormones are extracted from plasma with varying efficiency. Thus, markers or internal standards are often needed, to monitor and correct for extraction losses. To do so is difficult in the case of peptide hormones because radioactive recovery markers either have a low specific activity or, if labeled with iodine, may not be fully representative because of alterations in their size and charge. More importantly, markers labeled with 125I can interact in, and thus compromise, the subsequent radioimmunoassay. AtT-20 mouse pituitary tumor cells, which can be stimulated to synthesize and secrete pro-opiomelanocortin peptides, can biosynthetically label beta-lipotropin (beta-LPH) with [35S]methionine. The labeled peptide, which is co-eluted with unlabeled beta-LPH in "high-performance" liquid chromatography, is fully immunoprecipitable and has a specific activity of 34 Ci/g. We use this labeled peptide to monitor the recovery of beta-LPH in silicic acid extraction from plasma. This peptide is an ideal marker of analytical recovery because it does not interfere in subsequent radioimmunoassays.