The formation of a horseradish peroxidase-testosterone conjugate for the enzyme-linked immunoassay of testosterone was investigated, using tritiated testosterone to follow the reaction. The formation of testosterone-3-(carboxymethyl) oxime-peroxidase by the mixed anhydride method was found to give a conjugate of high enzymatic activity and with three molecules of testosterone per molecule of peroxidase. The optimum conditions for the assay of peroxidase activity were studied and an assay capable of measuring 1 to 5 ng of the conjugate developed; the standard curve being virtually linear. The stability of the conjugate in solution and the effect of lyophilisation on enzymatic activity are also described. The peroxidase-testosterone conjugate was suitable for enzyme-linked immunoassay and the quantities measurable with the peroxidase assay covered the range necessary for a plasma testosterone assay. The stability of the conjugate was such that no particular precautions were necessary for its storage.
Unexpected testosterone, LH, FSH and SHBG levels can result from illicit use of androgens (“anabolic steroids”). Classic symptoms include truncal acne, excessive muscularity, testicular atrophy and gynaecomastia, usually in association with obsessive and intense exercise regimens. Consider asking specific questions regarding androgen misuse to males who display these behaviours and signs. Males who are currently using androgens may have elevated testosterone and suppressed LH, FSH and SHBG. Suppression of testosterone as well as LH, FSH and SHBG can indicate a recent history of androgen misuse. 4 The rate of recovery from HPG axis suppression is dependent on the duration and severity of the misuse, but will generally occur within 12 months after cessation of androgens. 4
Bilirubin is a breakdown product of hemoglobin. Red blood cells contain hemoglobin, which is broken down into the components "heme" and "globin." Heme is converted into bilirubin. Albumin carries bilirubin to the liver, where most of it is conjugated with a glucuronide before it is excreted in the bile. Conjugated bilirubin is called "direct bilirubin;" unconjugated bilirubin is called "indirect bilirubin." Total bilirubin equals direct bilirubin plus indirect bilirubin.