|Andreotti, Peter - ATLANTIC SCIENTIFIC|
|Akhavan-Tafti, Hashem - LUMIGEN, INC.|
|Handley, Richard - LUMIGEN, INC.|
Submitted to: Journal of Immunological Methods
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: July 14, 2004
Publication Date: August 21, 2004
Citation: Gehring, A.G., Irwin, P.L., Reed, S.A., Tu, S., Andreotti, P.E., Akhavan-Tafti, H., Handley, R.S. 2004. Enzyme-linked immunomagnetic chemiluminescent detection of escherichia coli 0157:h7. Journal of Immunological Methods 293. 2004. p. 97-106. Interpretive Summary: Escherichia coli (E. coli) O157:H7 is a harmful bacterium that has been implicated in numerous cases of foodborne poisonings. A variety of rapid tests, that often require only a few hours for completion, have been created to detect the presence of this bacterium in foods. These screening tests are often used as alternatives to more conventional microbiological methods that often take days for an answer. Recent advances in light-measuring instrumentation (luminometers) have made the equipment more affordable and flexible in usage (i.e., portability for field testing). In an attempt to create better tests, we have developed a rapid test termed enzyme-linked immunomagnetic chemiluminescence (ELIMCL). ELIMCL incorporates this new generation of luminometers, antibodies (bound to very small magnetic particles) that selectively attach (and isolate bacteria from food samples) to E. coli O157:H7 bacterial cells, and amplified production of light as affected by the reaction of enzymes with added chemicals. This work presents the application of ELIMCL to the detection of E. coli O157:H7 with a detection limit of approx. 5,000 cells live cells in a buffered liquid. ELIMCL is also demonstrated to detect 10 bacterial cells of E. coli O157:H7 per gram of artificially contaminated ground beef in a total testing time of about 7 hours. ELIMCL may be used by food producers to test for the presence of harmful bacteria in their products.
Technical Abstract: E. coli O157:H7 is a pathogenic microorganism that has been implicated in numerous cases of foodborne illnesses. A variety of rapid methods exist that show promise for the presumptive detection of this pathogen without the immediate need for incubating test samples for hours to days in microbial enrichment media. Highly sensitive chemiluminescence has become a more affordable and portable detection method. We have developed a rapid method, termed enzyme-linked immunomagnetic chemiluminescence (ELIMCL) that incorporates the selectivity of antibodies with magnetic particle isolation, the signal amplification indicative of enzymes, and highly sensitive chemiluminescent detection. This work presents the application of ELIMCL to the detection of E. coli O157:H7 in pristine buffered saline with detection limits of approx. 1 e3 to 1 e4 of live cells. The blocking agent casein and the surfactant Tween-20 were used to lower background luminescence and thus maximize signal to noise ratios. ELIMCL was demonstrated to detect E. coli O157:H7 inoculated in ground beef at 10 cfu/g after a 5.5 hr enrichment and about 1.5 hr assay time for a total detection time of about 7 hr.