Comparison of Antibody-Direct Epifluorescent Filter Technique with the Most Probable Number Procedure for Rapid Enumeration of Listeria in Fresh Vegetables

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Abstract

The antibody-direct epifluorescent filter (Ab-DEFT) technique was evaluated as a rapid alternative to the most probable number (MPN) method for enumeration of artificially inoculated Listeria monocytogenes in ready-to-eat packaged salads and other fresh vegetables. Ab-DEFT was performed by homogenization of food in mesh-lined Stomacher bags, followed by prefiltration of homogenate through a 5 μm pore nylon filter, and passage of filtrate through a 0.4 μm pore black polycarbonate filter to collect and concentrate Listeria cells. After cells were stained with a fluorochrome-labeled polyclonal antibody to Listeria, the filter surface was examined by epifluorescence microscopy, and fluorescent cells were counted. A 3-tube MPN procedure was performed by successive enrichments of homogenized foods in Listeria enrichment and Fraser broths, followed by selective plating. Ab-DEFT provided quantitative determinations of Listeria cells that correlated with plate counts and MPN estimates in a linear response over a range of cell concentrations from 10 to 107 colony forming units (CFU)/mL. Microbial backgrounds as high as 108 CFU/mL did not affect performance of Ab-DEFT. In contrast to the MPN method, which required 5 days to perform, quantitation by Ab-DEFT could be completed in less than 1 h. Despite cross-reactivities demonstrated by the polyclonal fluorescent antibody, the potential of Ab-DEFT as a rapid alternative to MPN for microbial cell enumeration was evident.

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Tortorello, M. L., Reineke, K. F., & Stewart, D. S. (1997). Comparison of Antibody-Direct Epifluorescent Filter Technique with the Most Probable Number Procedure for Rapid Enumeration of Listeria in Fresh Vegetables. Journal of AOAC International, 80(6), 1208–1214. https://doi.org/10.1093/jaoac/80.6.1208

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