A radioimmunoassay of plasma ACTH, useful for several mammalian species, has been adapted for the rat. The antibody permitting this versatility was produced in the rabbit by immunization with the species-common, steroidogenic 1—24 amino acid sequence of ACTH. Binding studies, using polypeptide fragments of ACTH, showed that the antibody bound most effectively with the 1—24 fragment and nearly as effectively with the 1—39 native human and porcine molecules. The antibody failed to react significantly with αMSH, βP-MSH or with synthetic 1—16 amide or αp 17—39 fragments of ACTH. A close similarity was found between bioactivity and immunoreactivity in 3 tested specimens of rat plasma. Physiological validation of the method was obtained from the following studies. The immunoreactive plasma ACTH concentration at 9 AM in resting male rats was 23±4 pg/ml (mean±SE), when the plasma corticosterone concentration was 4±0.4 μg/100 ml. At 4:30 PM, plasma ACTH was 63±9 pg/ml and plasma corticosterone was 13±2 μg/100 ml. Intact male rats had elevations of immunoreactive plasma ACTH to 252—1910 pg/ml 2.5 min after the onset of ether stress. Adrenalectomized resting rats had elevated levels 24 hr, 7 days and 60 days post adrenalectomy. Their values were 195±22, 277±87 and 364±75 pg/ml, respectively. Ether-stressed rats, 24 hr, 7 days and 60 days after adrenalectomy had levels of 1010±232, 3375±340 and 7325±1103 pg/ml, respectively.
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