title: Three methods compared for estimating the fraction of testosterone and estradiol not bound to sex-hormone-binding globulin.
abstract: Here we propose and validate, both theoretically and experimentally, a new technique for measuring the percentage of non-sex-hormone-binding globulin (SHBG) bound to testosterone and estradiol. In this method, SHBG is saturated with 1 mumol of 5 alpha-dihydrotestosterone per liter and the non-protein-bound fraction is subsequently measured in treated and untreated samples by centrifugal ultrafiltration dialysis (CUFD). We compared this technique with two previously published methods for measuring the percentage of albumin-bound steroid--ammonium sulfate precipitation and heat denaturation of SHBG followed by CUFD--and also with previously published computer predictions. The ammonium sulfate precipitation gave results closest to the computer-predicted percentage of albumin-bound testosterone and estradiol. Of the two techniques involving CUFD, the 5 alpha-dihydrotestosterone saturation method was the more reproducible. Results by both CUFD methods were higher than the computer-based predictions for albumin-bound steroid in pregnancy serum, in part perhaps because the CUFD measurements represent the non-SHBG-bound fraction rather than that which is albumin-bound alone.
authors.affiliation: Department of Biochemical Endocrinology, Royal Marsden Hospital, London, U.K.
authors.forename: S
authors.initials: S
authors.lastname: Pearce
authors.forename: M
authors.initials: M
authors.lastname: Dowsett
authors.forename: S L
authors.initials: SL
authors.lastname: Jeffcoate
mesh_terms: D000645:Ammonium Sulfate; D002498:Centrifugation; D011232:Chemical Precipitation; D003956:Dialysis; D013196:Dihydrotestosterone / Q000378:metabolism; D004958:Estradiol / Q000097:blood*; D005260:Female; D006358:Hot Temperature; D006801:Humans; D007700:Kinetics; D008297:Male; D011247:Pregnancy; D011489:Protein Denaturation; D012709:Serum Albumin / Q000378:metabolism; D012738:Sex Hormone-Binding Globulin / Q000378:metabolism*; D013739:Testosterone / Q000097:blood*; D014462:Ultrafiltration
chemical_list: D012709:Serum Albumin; D012738:Sex Hormone-Binding Globulin; D013196:Dihydrotestosterone; D013739:Testosterone; D004958:Estradiol; D000645:Ammonium Sulfate