A Specific Radioimmunoassay of a hCG-β Subunit in the Presence of Native hCG, and Immunological, Biological and Chemical Characterization of hCG and its Chemically Modified Subunits

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  • hCGおよびhCG-βsubunitの免疫化学的, 生物学的, 化学的研究
  • hCGおよびhG-β subunitの免疫化学的,生物学的,化学的研究
  • hCG オヨビ hG ベータ subunit ノ メンエキ カガクテキ セイブ

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We have succeeded in preparing a specific radioimmunoassay (RIA) of a hCG-β subunit independent of native hCG by using a specific antiserum of the hCG-β subunit. The antiserum obtained by an absorption procedure was specific enough to assay the hCG-β subunit in serum and urinary specimens collected from pregnant women and a subject with a cancerous disease. A rabbit antibody against the conformational specificity of antigenic determinants of the hCG-β subunit was obtained by the following procedures : <BR>(1) A chemically purified and fractionated hCG-β subunit was first purified by an absorption technique using anti-normal human serum and anti-urinary Albert's B fraction antisera.<BR>(2) The antigen was then absorbed and purified by an anti-hCG antiserum which has a higher association constant (Ka) for antigenic determinant of native hCG and a lower Ka for the hCG-β subunit.<BR>(3) The antiserum was obtained by multiple intradermal injections of about 100 μg of the antigen thus purified.<BR>(4) The antibody against native hCG in the anti-hCG-β subunit antiserum with higher Ka for the hCG-β subunit was neutralized with highly purified parent hCG.<BR>The cross reaction between the anti-hCG-β subunit antisera thus purified and hCG was only from 2.7 to 3.3%, whereas with the anti-hCG-β subunit antisera without the neutralization procedure, there was from 6.3 to 11.0% by Ka calculation or 50% binding in RIA. The peak levels of the hCG-β subunit in the urine were obtained at the 11th-13th weeks after pregnancy, whereas the peak did not appear in the serum specimens in the purified specific RIA. The urine from a patient with gastric carcinoma was characterized by a gradual elevation of specific hCG-β subunits in contrast with minor changes of antigens determined by the conventional RIA, the values of which were possibly under the influence of native hCG levels.<BR>We also studied the chemical and enzymatic effect of hCG and its α and β subunits on immunological, biological and chemical behavior. Using citraconic and maleic acids for the modification of hCG and its subunits, we confirmed that lysine residues in the hCG-α subunit and arginine residues in the hCG-β subunit were strongly responsible for biological activity. The plasmin digestion of hCG for 3 hr gave a strong formation of immune complex with the anti-hCG-β subunit antiserum and for 8 hr formed a strong precipitin line with antiserum to the hCG-α subunit by gel diffusion analysis. These results show further possibilities for developing specific assays for hCG subunits.

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