Development of a Novel Protein Chip-Based Test for Rapid and Cost-effective Salmonella Serotyping

Salmonella is one of the most important agents of food-borne illnesses. Serotyping is the most important typing method for characterization of Salmonella isolates, and involves the immunological classification of two surface structures, lipopolysaccharide (O antigens) and Phase 1 and Phase 2 flagella antigens, based on the Kauffmann-White scheme. This scheme employs more than 250 antisera for identification of over 2500 Salmonella serotypes. The current Salmonella serotyping method does not allow detection of multiple antigens simultaneously, and requires a minimum of three antibody-antigen reactions to identify a particular serovar. It takes at least two to three days to provide an answer, and is available mainly in reference laboratories. The objective of this study is to develop a rapid, cost-effective and easy-to-use alternative assay for Salmonella serotyping based on the protein microarray technology.
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In this project, methods have been established to immobilize Salmonella antibodies onto the glass slides, to label Salmonella cells directly with a fluorescent dye (Eosin Y or Cy3), and to capture and specifically detect the labeled Salmonella cells with the antibody array. A model antibody (6x6) array was constructed for identification of Salmonella serotypes: Enteritidis, Heildelberg and Typhimurium. The model array was able to detect Salmonella O antigens, and Phase 1 and Phase 2 flagella antigens simultaneously, and correctly identify the three serotypes when it was tested using 10 Salmonella reference strains. The model array has been expanded for typing 20 commonly-identified and clinically-important Salmonella serotypes. The expanded array is being optimized. The specificity and sensitivity of the array will be determined using a limited number of Salmonella and non-Salmonella bacteria, and verified with the standard Kauffmann-White typing scheme.