Measurement of Multiple Forms of Human Growth Hormone: Cross-Reactivities in Conventional and Two-Chain Radioimmunoassays*

Abstract

A RIA system to measure two-chain forms of human GH (hGH) is described in this report. The need for such a RIA became evident with the identification of several single and two-chain peptides separated from pituitary hGH. hGH-S, a mixture of three peptides (des 140–149 and des 140–146 with residue 152 either Asn or Asp) was produced by subtilisin cleavage of hGH and provided a basis for development of the RIA of two-chain forms of hGH; in this RIA, two-chain forms were used as antigen, for iodination, and for standards. Competitive binding-displacement characteristics of several naturally occurring single chain and two-chain forms of hGH were assessed in the hGH-S RIA system and in a standardized hGH RIA using materials supplied by NIAMDD. In addition, enzymically produced two-chain forms were tested. These studies show that the antiserum used in the hGH RIA reacts to varying degrees with both single and two-chain forms of GH, whereas the antiserum to hGH-S reacts only with some forms of two-chain hGH and their N-terminal fragments (F₁), obtained by reduction of the disulfide bridges, followed by alkylation of the two-chain forms of hGH. The hGH-S RIA may be useful for the measurement of twochain forms of hGH found in pituitary extracts as well as in blood. Specific assays such as this may provide a means to resolve some of the contradictory data seen when results from RIAs are compared with results from either bioassays or radioreceptor assays. (Endocrinology106: 92, 1980)