Ultrasensitive luminol electrochemiluminescence for protein detection based on in situ generated hydrogen peroxide as coreactant with glucose oxidase anchored AuNPs@MWCNTs labeling

Abstract

In this study, an ultrasensitive luminol electrochemiluminescence (ECL) immunosensor was constructed using carboxyl group functionalized multi-walled carbon nanotubes (MWCNTs) as platform and glucose oxidase (GOD) supported on Au nanoparticles (AuNPs) decorated MWCNTs (AuNPs@MWCNTs–GOD) as labels. Firstly, using poly(ethylenimine) (PEI) as linkage reagents, AuNPs@MWCNTs were prepared and introduced for binding of the secondary antibody (Ab2) and glucose oxidase (GOD) with high loading amount and good biological activity due to the improved surface area of AuNPs@MWCNTs and excellent biocompatibility of AuNPs. Then the GOD and Ab2 labeled AuNPs@MWCNTs were linked to the electrode surface via sandwich immunoreactions. These localized GOD and AuNPs amplified luminol ECL signals dramatically, which was achieved by efficient catalysis of the GOD and AuNPs towards the oxidation of glucose to in situ generate improved amount of hydrogen peroxide (H2O2) as coreactant and the enhancement of AuNPs to the ECL reaction of luminol–H2O2. The experimental results demonstrated that the proposed immunosensor exhibited sensitive and stable response for the detection of α-1-fetoprotein (AFP), ranging from 0.0001 to 80ngmL−1 with a limit of detection down to 0.03pgmL−1 (S/N=3). With excellent stability, sensitivity, selectivity and simplicity, the proposed luminol ECL immunosensor showed great potential in clinical applications.