牙本质涎磷蛋白电化学酶联免疫检测方法初探

Sensitive detection of dentin sialophosphoprotein by electrochemical enzyme-linked immunoassay

目的初探一种检测正畸牙根吸收特异性蛋白的新型电化学酶联免疫分析(ELISA)方法。方法选取人牙本质涎磷蛋白(DSPP)标准品以辣根过氧化物酶(HRP)为标记酶、四邻联甲苯胺(OT)为酶催化反应的底物,分别用电化学ELISA法及传统光学ELISA法检测酶催化产物。对比研究两种检测方法下DSPP检测的线性范围及检测限的差异。结果用电化学ELISA法检测酶催化产物,产物在-0.63 V(vs.Ag/Ag Cl)有一个灵敏的还原峰,进而可以用于游离HRP的检测。应用于DSPP标准品的检测,线性范围为0.5-800.0 pg/m L,检出限为0.5 pg/m L,是传统光度ELISA检测限的1/10。结论电化学ELISA法可能成为检测牙根吸收特异性蛋白的新型生化检测方法。

Objective To investigate the electrochemical enzyme-linked immunoassay for the detection of dentin sialophosphoprotein （DSPP）. Methods Standard DSPP was detected by electrochemical enzyme-linked immunoassay and traditional spectroscopic enzyme-linked immunosorbent assay （ELISA） respectively. The different resuhs of the two methods were analyzed. Results HRP（ O-tilidine-H2O2- horseradish peroxidase）could catalyze the oxidation of OT with H2O2 to produce an electroactive product, which exhibited a sensitive second order derivative linear sweep polarographic reduction peak at -0.63 V （vs. Ag/AgC1）. By combined with the sandwich ELISA procedure, DSPP could be further detected in the concentration range from 0.5 pg/mL to 800.0 pg/mL with the detection limit at 0.5 pg/mL, which was 10 times lower than spectroscopic ELISA method. Conclusion Electrochemical enzyme-linked immunoassay is a new way of detecting DSPP.