Chemiluminescence assay of glucose oxidase using lucigenin-copper(II)ion and its application to enzyme immunoassay.

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  • ルシゲニンと銅（ＩＩ）イオンを用いたグルコースオキシダーゼの化学発光測定法及びその酵素免疫測定法への応用
  • ルシゲニン ト ドウ 2 イオン オ モチイタ グルコースオキシダーゼ ノ カ

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Abstract

Lucigenin has not been applicable to glucose oxidase (GOD) determination because it reacts with both the substrate and product. We have investigated the effects of some metal ions on the luminescence reaction of lucigenin. Copper or manganese ions were added to the basic luminescence solution containing lucigenin, which was then mixed with a glucose or hydrogen peroxide solution. Luminescence intensity was monitorred for 060 s after mixing. The luminescence intensity of the lucigenin-glucose reaction was selectively decreased to 1.63.1% by adding copper or manganese ions; that of the lucigenin-hydrogen peroxide reaction was kept at the control level. By this selective effect, lucigenin could be used for GOD determination and for enzyme immunoassay with GOD. The assay procedure for GOD is as follows: 25 μl of GOD solution (10 fM-20 nM GOD in 20 mM sodium-acetate buffer pH 5.0 containing 10 mg/ml Triton X-100) was added to 25 μl of glucose solution (60 mM glucose in 20 mM sodium-acetate buffer pH 5.0) and then incubated at 4°C overnight. Five hundred microliters of luminescence solution (4× 10 -3% lucigenin, 25 mM sodium hydroxide, 100 mM sodium phosphate, dibasic, 0.1 % Triton X-100) was added and then the intensity of chemiluminescence was integrated. The detection limit for GOD with this system was 2.5 × 10 -9 mol/assay. The assays for estrogen and human chorionic gonadotropin (hCG) were examined as models for enzyme immunoassay with GOD as the labeled enzyme. When the estrogen and the hCG contents of urine samples were measured by this method and LPIA latex photometric immunoassay or RIA, the RSD were 0.972 and 0.975, respectively.

Journal

  • BUNSEKI KAGAKU

    BUNSEKI KAGAKU 41 (10), 473-478, 1992

    The Japan Society for Analytical Chemistry

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