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Antigenic complexes of 2-hydroxyestrone (2-OHE1) were obtained by Mannich reaction of 2-OHE1 and bovine serum albumin (BSA) and by coupling of 2-OHE1 1-glutathione thioether to BSA using glutaraldehyde. Antiserum raised against the antigen obtained by the Mannich reaction had high affinity (Kd = 3.8 x 10(9) M-1) and relatively high specificity; cross reactivities for estrone, 4-hydroxyestrone and 2-methoxyestrone were 2.1%, 10% and 1.5%, respectively. The other antiserum also had high affinity (4.5 x 10(9) M-1) but its cross reactivities for the above three steroids were more than 100%. Concentrations of 2-hydroxyestrone in human plasma were determined by radioimmunoassay with the more specific antiserum and Sephadex LH-20 chromatography to be less than a minimum detectable amount (less than 10 pg/ml) (men), 20.9 pg/ml (women, proliferation) and 26.0 pg/ml (women, periovulation).

Radioimmunoassay of 2-hydroxyestrone using antisera raised against antigenic complexes obtained by convenient methods.