A sandwich protocol for chemiluminescent detection of Streptococcus mutans utilizing dual-site recognition by rat IgG2a and vancomycin

Abstract

A sandwich protocol was developed for chemiluminescent detection of Streptococcus mutans (S. mutans) utilizing dual-site recognition of the bacteria by rat IgG2a and vancomycin. Magnetic particles (MPs) functionalized with rat IgG2a were adopted to capture S. mutans based on the binding between Fc region of rat IgG2a and protein G in the cell wall of the bacteria. Alkaline phosphatase labeled vancomycin was applied as the signal tracer since D-Ala-d-Ala peptide moieties in the cell wall of Gram-positive bacteria could be recognized by this antibiotic agent. The protocol showed an ideal specificity for S. mutans detection because the recognition agents bound with the bacteria at two distinct sites. Furthermore, the utilization of MPs enrichment greatly improved the sensitivity. S. mutans could be detected within a linear range of 2.4×101 − 2.4×105 CFU mL−1, with a very low detection limit of 2.3×101 CFU mL−1. Some common Gram-positive bacteria and Gram-negative bacteria showed negligible interference. This proposed method for S. mutans detection possessed some attractive characteristics such as facile manipulation, short detection time, low cost, ultra-high sensitivity and ideal specificity. It was used to detect S. mutans in human saliva with acceptable recoveries ranging from 70.8% to 108.3%.