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United States Department of Agriculture

Agricultural Research Service

Research Project: DETECTION AND TYPING OF FOOD-BORNE PATHOGENS

Location: Molecular Characterization of Foodborne Pathogens

Title: A high-throughput antibody-based microarray typing platform

Authors
item Gehring, Andrew
item Barnett, Charles -
item Chu, Ted
item Debroy, Chitrita -
item D'Souza, Doris -
item Eaker, Shannon -
item Fratamico, Pina
item Gillespie, Barabra -
item Hedge, Narasimha -
item Jones, Kevin -
item Lin, Jun -
item Oliver, Stephen -
item Paoli, George
item Perera, Ashan -
item Uknalis, Joseph

Research conducted cooperatively with:
item

Submitted to: Sensors
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: April 15, 2013
Publication Date: May 3, 2013
Citation: Gehring, A.G., Barnett, C., Chu, T., Debroy, C., D'Souza, D., Eaker, S., Fratamico, P.M., Gillespie, B., Hedge, N., Jones, K., Lin, J., Oliver, S., Paoli, G., Perera, A., Uknalis, J. 2013. A high-throughput antibody-based microarray typing platform. Sensors. DOI: 10.3390/s130505737.

Interpretive Summary: There are many rapid tests that can be used to screen foods for the presence of harmful bacteria. Such detection tests yield timely (minutes to hours) results for identifying foods that are suspected to be contaminated. Fast tests that have the ability to not only detect, but also type (categorize or identify) bacteria via recognition with biological elements (e.g., antibodies or DNA probes) may facilitate epidemiological investigations that aim to trace back the contamination to its source. This manuscript introduces a novel antibody-based typing array that functions in a high throughput, multiwell plate assay that uses lasers for generating measureable fluorescence signals for captured bacteria pretreated with a dye or stain called Sybr Gold. Two sets of antibodies generated to specifically react with “Big Six” non-O157 Shiga toxin-producing Escherichia coli (STEC) bacteria were printed into multiwell plates, and the bacteria were added and subsequently detected and identified. The STEC antibody sets compared reasonably well, though improved recognition of the different bacterial types increased with higher levels of antibody. The favorable results indicated that with ideal sets of biorecognition elements, this high-throughput array platform can rapidly detect and be used to type and potentially detect bacteria derived from food samples within approximately one hour.

Technical Abstract: Many rapid methods have been developed for screening foods for the presence of pathogenic microorganisms. Rapid methods that have the ability to identify microorganisms via multiplexed immunological recognition have the potential for classification or typing microbial contaminants thus facilitating epidemiological investigations that aim to trace back the contamination to its source. This manuscript introduces a novel antibody-based typing array that functions as a high throughput, multiwell plate assay that uses laser-induced fluorescence based on the nucleic acid intercalating dye/stain Sybr Gold for detection of antibody-captured bacteria. Two sets of antibodies raised against the “Big Six” non-O157 Shiga toxin-producing E. coli (STEC) were array-printed into microtiter plates and enriched samples of bacteria were added and subsequently detected. The STEC antibody sets compared reasonably well, though specificity increased at higher antibody dilution levels. The favorable results indicated that with ideal sets of biorecognition elements (e.g., antibodies, aptamers, and nucleic acid probes), this high-throughput platform can be used to rapidly type microbial isolates derived from food samples within ca. 1 h of total assay time. It can also potentially be used to detect the pathogens from food enrichments.

Last Modified: 8/30/2014