A B S T R A C T Extracts of tumors from 32 patients with the ectopic ACTH syndrome were subjected to simultaneous bioassay and radioimmunoassays for ACTH. Radioimmunoassays were performed using three antisera, one of which reacts with the extreme N-terminal 1-13 amino acid sequence of ACTH, the second with the N-terminal 1-23 sequence of the ACTH molecule, and the third with the C-terminal 25-39 amino acid sequence of ACTH. There was, in general, good correlation between bioactivity and N-terminal ACTH immunoreactivity. However, there were large excesses of both extreme N-terminal and C-terminal immunoreactive materials in most tumor extracts, which were not found in extracts of three human pituitaries. Three tumor extracts were subjected to molecular sieve chromatography on Sephadex G-50 fine resin. The bioactive ACTH eluted in the same fractions as pituitary ACTH (mol wt