检测血清p185糖链凝集素酶联免疫吸附试验的建立及初步应用

Establishment and preliminary application of lectin enzyme-liked immunosorbentassay for serum p185 carbohydrate chain

目的建立检测血清p185糖链的凝集素酶联免疫吸附试验(ELISA),并初步应用于乳腺癌的临床诊断。方法用改良过碘酸钠氧化法制备c-erbB-2单克隆抗体-辣根过氧化物酶结合物(A18-HRP),经棋盘滴定确认A18-HRP、生物素化麦胚凝集素(B io-WGA)及链霉亲和素(SA)的最适浓度(或稀释度),建立了检测p185糖链的凝集素ELISA,用所建立的方法检测血清p185糖链,免疫组化方法检测组织p185蛋白表达。结果乳腺癌患者血清p185糖链水平明显高于乳腺增生和正常对照(P<0.01),免疫组化阳性和阴性的乳腺癌患者血清p185糖链水平均高于乳腺增生和正常对照(P<0.01),乳腺增生患者血清p185糖链与正常对照相比差异无显著性(P>0.05)。血清p185糖链诊断乳腺癌的敏感度为86.7%,特异度为94.6%,准确度为91.9%。结论本实验建立的凝集素ELISA检测血清p185糖链诊断乳腺癌与病理学诊断符合率高,临床具有实用价值。

Objective To establish a new lectin enzyme-liked immunosorbent assay （ELISA） for serum p185 carbohydrate chain and apply it in the diagnosis of the breast cancer. Methods A18-Cerb-2 monoclonal antibody conjugate （A18-HRP） was prepared with modified sodium heptaiodic acid oxidation method. Optical concentration of A18- HRP, biotingl wheat germ agglutinin （Bio-WGA） and streptavidin （SA） were affirmed by chess-board titration and lectin ELISA serum p185 carbohydrate chain was established. Serum p185 carbohydrate chain was detected by lectin ELISA; p185 protein expression in tissue was detected by immunohistochemical technique. Results The level of serum p185 carbohydrate chain in breast cancer was significantly higher than that in breast proliferation and normal control （P 〈 0.01 ）. The level of serum p185 carbohydrate chain in breast cancer with positive and negative tissue p185 expression was significantly higher than that in breast proliferation and normal control （P 〈0.01 ）. There was no obvious difference between breast proliferation and normal control （P 〉 0.05 ）. The sensitivity, specificity and accuracy of serum p185 carbohydrate chain in breast cancer were 86.7%, 94.6% and 91.9% respectively. Conclusions The lectin ELISA for serum p185 carbohydrate chain accords highly with pathologic diagnosis of breast cancer and has its value in clinical application.