An electrochemical immunoassay for Escherichia coli O157:H7 using double functionalized Au@Pt/SiO2 nanocomposites and immune magnetic nanoparticles

A sensitive Point-of-Care Testing (POCT) with Au-Pt bimetallic nanoparticles (Au@Pt) functionalized silica nanoparticle (SiO2 NPs) and Fe3O4 magnetic nanoparticles (Fe3O4 NPs) was designed for the quantitative detection of Escherichia coli O157:H7 (E. coli O157:H7). The poly-(4-styrenesulfonic acid-co-maleic acid) (PSSMA) as a negatively charged polyelectrolyte can be easily coated on surface of the amino group modified SiO2 NPs via electrostatic force. PSSMA is also a good stabilizer for water-soluble bimetallic nanostructures. The PSSMA is first time used as a bridge to connect the negative charge Au@Pt NPs to the SiO2 NPs, forming Au@Pt/SiO2 NPs. Antibody and invertase conjugated Au@Pt/SiO2 NPs (denoted as Ab/invertase-Au@Pt/SiO2 NPs) were used as signal labels. Monoclonal antibody against E. coli O157:H7 (Ab) functionalized magnetic nanoparticles (denoted as Ab-Fe3O4@SiO2 NPs) were used to enrich and capture the E. coli O157:H7 in positive sample. The immunosensing platform also composed of a personal glucometer (PGM) using for signal readout. Based on this sandwich-type immunoassay, the invertase in the final formed sandwich immunocomplex catalyzed the hydrolysis of sucrose to produce a large amount of glucose for quantitative readout by the PGM. Under optimal conditions, a linear relationship between the glucose concentration and the logarithm of E. coli O157:H7 concentration was obtained in the concentration range from 3.5 x 10(2) to 3.5 x 10(8) CFU mL(-1) with a detection limit of 1.83 x 10(2) CFU mL(-1) (3 sigma). This method was used to detect E. coli O157:H7 in spiked milk samples, indicating its potential practical application. This protocol can be applied in various fields of study.