Competitive binding of polyethyleneimine-coated gold nanoparticles to enzymes and bacteria: a key mechanism for low-level colorimetric detection of gram-positive and gram-negative bacteria

The article describes a simple and rapid method for colorimetric detection of bacteria. It is based on competitive binding of positively charged polyethyleneimine-coated gold nanoparticles (PEI-AuNPs) to negatively charged enzymes and bacteria. The PEI-AuNPs are electrostatically attracted by both the bacterial surface and the enzyme β-galactosidase (β-Gal). Binding to the latter results in the inhibition of enzyme activity. However, in the presence of a large number of bacteria, the PEI-AuNPs preferentially bind to bacteria. Hence, the enzyme will not be inhibited and its activity can be colorimetrically determined via hydrolysis of the chromogenic substrate chlorophenol red β-D-galactopyranoside (CPRG). The detection limit of this assay is as low as 10 cfu·mL −1 , and the linear range extends from 10 6 to 10 8 cfu·mL −1 . The assay is applicable to both Gram-negative (such as enterotoxigenic Escherichia coli ; ETEC) and Gram-positive ( Staphylococcus aureus ; S. aureus ) bacteria. Results are obtained within 10 min using an optical reader, and within 2–3 h by bare-eye detection. The method was applied to the identification of ETEC contamination at a level of 10 cfu·mL −1 in spiked drinking water. Given its low detection limit and rapidity (sample preconcentration is not required), this method holds great promise for on-site detection of total bacterial contamination. Graphical Abstract The method is based on competitive binding of positively charged polyethyleneimine-coated gold nanoparticles to negatively charged enzymes and bacteria. The detection limit is as low as 10 cfu·mL −1 , and the linear range extends from 10 6 to 10 8 cfu·mL −1 .