This Article Full Text Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Reprints and Permissions Copyright Information Books from ASM Press MicrobeWorld Citing Articles Citing Articles via Google Scholar Google Scholar Articles by DeCory, T. R. Articles by Montagna, R. A. Search for Related Content PubMed PubMed Citation Articles by DeCory, T. R. Articles by Montagna, R. A. Agricola Articles by DeCory, T. R. Articles by Montagna, R. A.

 Previous Article  |  Next Article 

Applied and Environmental Microbiology, April 2005, p. 1856-1864, Vol. 71, No. 4
0099-2240/05/$08.00+0     doi:10.1128/AEM.71.4.1856-1864.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.

Development of an Immunomagnetic Bead-Immunoliposome Fluorescence Assay for Rapid Detection of Escherichia coli O157:H7 in Aqueous Samples and Comparison of the Assay with a Standard Microbiological Method

Thomas R. DeCory,¹ Richard A. Durst,¹ Scott J. Zimmerman,^2,3 Linda A. Garringer,² Gary Paluca,² Heleen H. DeCory,^1, and Richard A. Montagna^4*

Department of Food Science and Technology, Cornell University, Geneva,¹ Clinical Center, Public Health Laboratory, Department of Health, County of Erie,² Department of Microbiology, School of Medicine, and School of Public Health and Related Professions, State University of New York at Buffalo, Buffalo,³ Innovative Biotechnologies International, Inc., Grand Island, New York⁴

Received 8 August 2004/ Accepted 1 November 2004

The objective of this study was to develop and optimize a protocol for the rapid detection of Escherichia coli O157:H7 in aqueous samples by a combined immunomagnetic bead-immunoliposome (IMB/IL) fluorescence assay. The protocol consisted of the filtration or centrifugation of 30- to 100-ml samples followed by incubation of the filter membranes or pellet with anti-E. coli O157:H7 immunomagnetic beads in growth medium specific for E. coli O157:H7. The resulting E. coli O157:H7-immunomagnetic bead complexes were isolated by magnetic separation, washed, and incubated with sulforhodamine B-containing immunoliposomes specific for E. coli O157:H7; the final immunomagnetic bead-E. coli O157:H7-immunoliposome complexes were again isolated by magnetic separation, washed, and lysed with a n-octyl-ß-D-glucopyranoside to release sulforhodamine B. The final protocol took less than 8 h to complete and had a detection limit of less than 1 CFU of E. coli O157:H7 per ml in various aqueous matrices, including apple juice and cider. To validate the protocol at an independent facility, 100-ml samples of groundwater with and without E. coli O157:H7 (15 CFU) were analyzed by a public health laboratory using the optimized protocol and a standard microbiological method. While the IMB/IL fluorescence assay was able to identify E. coli O157:H7-containing samples with 100% accuracy, the standard microbiological method was unable to distinguish E. coli O157:H7-spiked samples from negative controls without further extensive workup. These results demonstrate the feasibility of using immunomagnetic beads in combination with sulforhodamine B-encapsulating immunoliposomes for the rapid detection of E. coli O157:H7 in aqueous samples.

Present address: Celltech Americas, Inc., Rochester, NY 14623.