血清CBGと特異的に結合する免疫吸着剤の作成と, それを用いたCBGとcortisolの相互作用の研究:健常者でのCBGとcortisolのKa値の検討

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We have developed a simple method for the evaluation of the association constant (Ka) between cortisol binding globulin (CBG) and cortisol using immunoadsorbent and a small amount of serum. Immunoadsorbent was prepared as follows; gamma-globulin fraction was obtained from goat anti-human CBG antiserum by salting out using 50% ammonium sulfate, followed by coupling to CNBr-activated Sepharose 4B. Immunoadsorbent thus prepared was used for the evaluation of Ka value between cortisol and CBG. Sera obtained from 5 healthy subjects (3 males and 2 females) were treated with charcoal (Norit A) to remove endogeneous cortisol. Each of 10μl of serum was incubated with a constant amount of 125I-cortisol and different concentrations of unlabelled cortisol for 12 hours at 25°C, pH 7.4. Then, 50μlf anti-CBG-Sepharose 4B (immunoadsorbent) was added to each incubation mixture, followed by incubation for another 4 hours at 25°C. After centrifugation, radioactivity of the precipitate was counted and the results were analyzed using Scatchard plot. The Ka value between serum CBG and cortisol in 5 healthy subjects was 1.70 × 108M-1 (SD = 0.24), which is comparable with that of a previously reported value using purified CBG from a large amount of serum. Interassay variation of this method was satisfactory with CV value of 10.1%. Our present method is considered to be clinically relevant, since it does not require purification of CBG from serum, and a relatively small amount (total amount, less than 200μl) of serum is enough to evaluate Ka value between CBG and cortisol.
一般社団法人日本内分泌学会の論文

血清CBGと特異的に結合する免疫吸着剤の作成と, それを用いたCBGとcortisolの相互作用の研究:健常者でのCBGとcortisolのKa値の検討

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