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 beta-galactosidase (beta-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 beta-D-galactopyranoside (CPRG). The detection limit of this assay is as low as 10 cfu center dot mL(-1), and the linear range extends from 10(6) to 10(8) cfu center dot 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 center dot 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.