以邻苯二胺为底物固体电极方波伏安法检测辣根过氧化物酶及其标记物

Square Wave Voltammetric Determination of Horseradish Peroxidase and Enzyme Labeled IgG Based on o-phenylenediamine as Substrate at Glassy Carbon Electrode

以玻碳电极为工作电极,研究了邻苯二胺(OPD)为底物伏安法检测辣根过氧化物酶(HRP)及其标记物的方法。HRP能够催化H_2O_2氧化OPD,其反应产物在玻碳电极上于-0.42V(vs.Ag/Agcl)左右产生一个灵敏的还原峰,峰电流随着HRP浓度的增大而增大,借助此还原电流可以测定HRP,并进而可用于以HRP为标记物的电化学酶免疫分析。用方波伏安法对酶催化反应条件和酶催化反应产物的测定条件进行了详细的研究,在最佳条件下测定游离HRP的线性范围是1.0×10^(-10)～4.0×10^(-9)g/mL,检出限为8.5×10^(-11)g/mL;对游离的酶标记物(IgG-HRP)的测定最大稀释比为1:2000000。

A voltammetric method for the detection of horseradish peroxidase (HRP) and enzyme labeled IgG at glassy carbon electrode(GCE) was investigated based on o-phenylenediamine(OPD)-H_2O_2-HRP enzymatic reaction system. HRP can catalyze the reaction of H_2O_2 oxidizing OPD. The product was reduced at —0.42 V (vs. Ag/AgCl) at glassy carbon electrode and square wave vohammetry (SWV) was chosen to detect the enzymatic-generated product. The conditions for HRP enzymatic reaction and electrochemical detection were carefully studied with a CHI 832 electrochemical analyzer. The reduetive peak current was linear with the HRP concentration from 1.0×10^(-10) to 4.0×10^(-9)g/mL and the detection limit for HRP was 8.5×10^(-11)g/mL. The highest dilution ratio of free IgG-HRP was 1:2000000. This system can be further used in HRP-labeled electrochemical enzyme-linked immunoassay.