Optimized dendrimer-encapsulated gold nanoparticles and enhanced carbon nanotube nanoprobes for amplified electrochemical immunoassay of E. coli in dairy product based on enzymatically induced deposition of polyaniline

A highly sensitive immunosensor was reported for Escherichia coli assay in dairy product based on electrochemical measurement of polyaniline (PAn) that was catalytically deposited by horseradish peroxidase (HRP) labels. Herein, the immunosensor was developed by using poly(amidoamine) dendrimer-encapsulated gold nanoparticles (PAMAM(Au)) as sensing platform. Importantly, the optimal HAuCl4/PAMAM ratio was investigated to design the efficient PAMAM(Au) nanocomposites. The nanocomposites were proven to not only increase the amount of immobilized capture antibody (cAb), but also accelerate the electron transfer process. Moreover, the {dAb-CNT-HRP} nanoprobes were prepared by exploiting the amplification effect of multiwalled carbon nanotubes (CNTs) for loading detection antibody (dAb) and enormous HRP labels. After a sandwich immunoreaction, the quantitatively captured nanoprobes could catalyze oxidation aniline to produce electroactive PAn for electrochemical measurement. On the basis of signal amplification of the PAMAM(Au)-based immunosensor and the {dAb-CNT-HRP} nanoprobes, the proposed strategy exhibited a linear relationship between the peak current of PAn and the logarithmic value of E. coli concentration ranging from 1.0×102 to 1.0×106 cfu mL−1 with a detection limit of 50 cfu mL−1 (S/N=3), and the electrochemical detection of E. coli could be achieved in 3 h. The electrochemical immunosensor was also used to determine E. coli in dairy product (pure fresh milk, infant milk powder, yogurt in shelf-life and expired yogurt), and the recoveries of standard additions were in the range of 96.8-108.7%. Overall, this method gave a useful protocol for E. coli assay with high sensitivity, acceptable accuracy and satisfying stability, and thus provided a powerful tool to estimate the quality of dairy product.