A Comparison of Procedures for the Isolation of Adrenocorticotrophin From Human Pituitary Glands

Abstract

SUMMARY Two methods for the purification of corticotrophin from human pituitary glands have been assessed by two radioimmunoassays directed towards the C-terminal and N-terminal parts of the molecule, and by bioassay. One procedure, applied to small amounts of tissue, employs acid acetone extraction, oxycellulose adsorption and chromatography on CM-cellulose (CM-23). It is suitable for the small-scale purification of corticotrophin from pituitary glands but the losses associated with extracting low concentrations of hormone suggest that it would be inadequate for application to extrapituitary tissues and tumours. The starting material for the second procedure was a side fraction derived from the large scale extraction of protein hormones from whole pituitary glands. The purification employs oxycellulose adsorption, gel filtration on Sephadex G-50 and chromatography on CM-cellulose (CM-32). It yields sufficient highly potent corticotrophin for use as radioimmunoassay standard while conserving other scarce human pituitary hormones.

Review Article: Acth and Related Peptides

“…Whenever measurement of biologically active ACTH, secretion dynamics or plasma half-life is important, an antiserum should be chosen which shows minimal reaction with non-biologically active fragments. It is also desirable to estimate both C-terminal and N-terminal immunoreactivity when studying the release of ACTH-like peptides from non-endocrine tumours and when monitoring the purity of pituitary extracts (Thomas et al, 1973). The extraction procedure used may also affect specificity, for it is our experience using a silicic acid absorption method that the relative recoveries of ACTH, 8and yLPH are affected by slight changes of methodology.…”