Testosterone (T) and estradiol (E₂) are bound to serum proteins, and it is believed that this binding affects the biological activity of these hormones. Testosterone-estradiol binding globulin (TeBG) is the primary β-globulin which binds both T and E₂ specifically.
In recent years, various assay methods for determining TeBG capacity (TeBGC) and %-bound T, E₂ to TeBG (%T-TeBG, %E₂-TeBG) have been developed but most of them too laborious and complex to apply in practice.
We have developed a rapid and simplified method to determine TeBGC and %T, E₂-TeBG by means of a modified ammonium sulfate precipitation method;
1) Assay method for TeBGC
(1) to a series of assay tubes, add ³ H-DHT and 5ng DHT (control; 500ng DHT)
(2) dry up under gentle N₂ gas stream at 40°C
(3) add 0.1 ml of undiluted serum (both male and female) or diluted serum (pregnant) to the appropriate tube, and allow to incubate at 37°C for 30 min.
(4) add 0.5 ml of ice-cold 60%-saturated ammonium sulfate
(5) keep in ice bath for 15 min.
(6) centrifuge at 1500 xg at 0°C for 30 min.
(7) count 0.2 ml supernatant and calculate
2) Assay method for %T, E₂-TeBG
(1) to a series of assay tubes, add ³ H-T or ³ H-E₂ (control; with 500 ng T or E₂) Steps (2), (3), (4), (5), (6) and (7) are processed as in the TeBGC method except using undiluted serum in every case.
The values obtained were 9.9 ± 3.4 (mean ± SD) for male (expressed as ng DHT/ml), 15.3 ± 3.9 for female and 87.1 ± 16.9 for pregnant. While %T-TeBG was 61.6 ± 5.1% for male, 76.8 ± 2.8% for female, and 92.4 ± 4.3% for pregnant, %E₂-TeBG was 25.0 ± 4.0% for male, 48.1 ± 3.4% for female and 80.3 ± 6.3% for pregnant.
The values thus obtained for %T-and %E₂-TeBG by the author's ammoniuum sulfate precipitation method are highly correlated with those which are determined by the equilibrium dialysis method. Correlation coefficients are 0.99 for %T-TeBG and 0.97 for %E₂-TeBG.