A simple, rapid, and precise direct radioimmunoassay method for 17 alpha, 20 beta-dihydroxy-4-pregnen-3-one in salmon...

This report describes a radioimmunoassay method for 17 alpha,20 beta-dihydroxy-4-pregnen-3-one (17 alpha,20 beta-diOHP) in the blood plasma of Atlantic salmon (Salmo salar). 3H-labeled 17 alpha,20 beta-diOHP was synthesized from 17 alpha-[3H]hydroxy-4-pregnen-3-one (17 alpha-OHP) by enzymatic reduction of the C-20 oxo group and the simultaneous oxidation of coenzyme-reduced nicotinamide adenine dinucleotide. Antiserum to 17 alpha,20 beta-diOHP-3-CMO-BSA exhibited high specificity: all steroids tested against the antiserum gave less than 1% cross-reactivity at 50% displacement. Crude dichloromethane extracts of small volumes (50 microliters or less) of plasma can be assayed directly without prior purification. The working range is 5 to 300 pg. Interassay and intraassay coefficients of variation were less than 10%. Various parallelism tests showed high accuracy and sensitivity. The assay is simple and rapid and at least 75 samples can be easily assayed in duplicate by two analysts in 1 day.