电化学酶联免疫分析法测定人牙本质涎磷蛋白的研究

摘要

目的 探讨电化学酶联免疫分析法(ELISA)测定牙本质涎磷蛋白(DSPP)的可行性.方法 选取牙本质涎磷蛋白标准品以辣根过氧化物酶(HRP)为标记酶、邻联茴香胺(ODA)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.对比两种检测方法对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/AgCl)有一个灵敏的还原峰,进而可以用于游离HRP的检测,其线性范围为0.04~1.0 ng/mL,检测限为0.01 ng/mL.应用于DSPP标准品的检测,线性范围为2.5~200.0 pg/mL,检出限为2.5 pg/mL,灵敏度显著高于传统光度ELISA检测法.结论 电化学ELISA法可以作为检测痕量DSPP的一个新方法.%Objective To investigate the sensitivity of electrochemical enzyme -linked immunoassay for the detection of dentin sialophosphoprotein ( DSPP) . Methods Standard DSPP were detected by electrochemical enzyme -linked immunoassay and traditional spectroscopic enzyme -linked immunosorbent assay ( ELISA) respectively. The sensitivity of the two methods was compared and analyzed. Results HRP could catalyze the oxidation of ODA to produce an electroactive product, which exhibited a reduction peak at -0. 63 V ( vs. Ag/AgQ ) . Free HRP was detected in the concentration range from 0. 04 to 1. 0 ng/mL, with the detection limit of 0. 01 ng/mL DSPP, in combination with the sandwich ELISA procedure , could be further detected in the concentration range from 2. 5 pg/mL to 200. 0 pg/mL with the detection limit of 2. 5 pg/mL. The sensitivity of electrochemical enzyme -linked immunoassay was significantly higher than that of traditional spectroscopic ELISA. Conclusion Electrochemical enzyme -linked immunoassay is a new way of detecting DSPP with more sensitivity.