A sensitive radioimmunoassay for corticotropin using a fully biologically active 125I-labeled ligand.
The human corticotropin (ACTH) analog, Phe2,Nle4-ACTH-(1-38) was iodinated by the chloramine-T procedure and the product was purified by reverse phase high performance liquid chromatography. The specific radioactivity of [125I]Tyr23,Phe2,Nle4-ACTH-(1-38) was determined by comparing the antiserum binding curves of the iodinated peptide and [3H]ACTH of known specific activity. This method gave a value of 1800 +/- 75 Ci/mmol, which is close to the theoretical radioactivity expected for the introduction of a single 125I atom into the peptide. [125I]Tyr23,Phe2,Nle4-ACTH-(1-38) was as potent as ACTH in stimulating corticosterone production in isolated rat adrenocortical cells. The concentrations for half-maximal steroidogenesis were 36.5 +/- 6.1 pM for the 125I derivative and 37.6 +/- 6.7 pM for ACTH. By the use of this 125I-labeled ligand, a highly sensitive RIA capable of detecting 1 pg aCTH was developed.l The antiserum employed in this study appeared to be directed against residues 11-13 of ACTH.
D000324:Adrenocorticotropic Hormone / Q000031:analogs & derivatives* / Q000032:analysis* / Q000302:isolation & purification; D000818:Animals; D006801:Humans; D007106:Immune Sera; D007457:Iodine Radioisotopes; D010446:Peptide Fragments; D011817:Rabbits / Q000276:immunology; D011863:Radioimmunoassay / Q000379:methods
D007106:Immune Sera; D007457:Iodine Radioisotopes; D010446:Peptide Fragments; C030702:ACTH (1-38), Phe(2)-Nle(4)-iodo-Tyr(23)-; D000324:Adrenocorticotropic Hormone