Rapid Detection of Enterotoxigenic Escherichia coli O6 in Water by Using Monoclonal Antibody and a Photon-Counting Television Camera

Abstract

Monoclonal antibodies (MAbs) raised against Escherichia coli O6:H16 were screened against 15 strains of E. coli and 19 non–E. coli bacteria. A MAb-luminescence assay using MAb-5.8, which shows no cross-reactions with non–E. coli bacteria, and a photon-counting television camera were developed for rapid enumeration of E. coli O6:H16 in water. The membrane filter that retained bacteria was boiled for 5 min in a buffer and incubated with biotinylated MAb-5.8. After incubation with streptavidin-peroxidase conjugate, it was reacted with luminol-based reaction mixture. Luminous image and light intensity of the filter was recorded with a Biocell Counter. Levels of E. coli O6 higher than 7 × 103 CFU were detected by the MAbluminescence assay when E. coli O6 was spotted onto the membrane filter. The sample that contained E. coli O6:H16 was filtered through a membrane filter, and the filter that retained bacteria was incubated on a filter paper soaked with nutrient broth supplemented with 0.5% NaCl at 37°C for 6 h. The number of light emission points on the filter correlated well with initial E. coli O6:H16 counts within the range of 1 to 3 × 102 CFU. The correlation coefficient was 0.89.