SBIR-STTR Award

Recent advances in biochemistry and immunology now make possible new approaches for detection and treatment of infectious diseases.The objective of this project is to develop a monoclonal antibody-based immunoassay for the Escherichia coli (E. coli) heat-stable enterotoxin (ST), which causes "traveler's diarrhea" in humans, and widespread, often fatal, infectious diarrhea (scours) in newborn livestock. Phase I goals will be (1) to generate a panel of murine monoclonal antibodies to the toxin, and (2) to commence development of a sensitive and versatile ST fluorescence immunoassay. Purified monoclonal antibodies will be conjugated to 0-galactosidase and tested in a heterogeneous competitive assay format. Investigators will pursue solid-phase methods for immobilization of ST-albui-nin conjugates, which will then compete with the test sample for binding to antibody-enzyme conjugates. Enzyme activity (and subsequently, ST concentrations) will be measured in a fluorescence assay, monitoring the formation of methyl-umbelliferone from the nonfluorescent substrate, met hyl-umbellif eryl galactoside. The final (fluorescence) measurements can be made in any appropriately set fluorimeter, or in a low-cost, portable photon-counting fluorimeter that investigators have been developing for use with such immunoassays. Both the instrument and the assay format are geared toward rapid, technically facile performance in a laboratory or "field" setting.Due to the health and economic impact of toxin-related diseases, the investigators believe there are significant human and veterinary commercial markets for such a toxin detection method, as well as the possible commercial utility of such antibodies as therapeutic agents. Genetics Diagnostics Corporation is also well positioned to attract follow-on funding for this project and to develop/market such commercial products. The technology developed should also be applicable to other infectious diseases.National Institute Of Allergy And Infectious Diseases

Genetic Diagnostics, Inc. is interested in pursuing the development of a rapid non-isotopic immunoassay for E. coli heat stable enterotoxin (Sta). Sta is produced by many pathogenic strains of E. coli, and causes acute, often fatal, diarrheal disease in humans and livestock animals.Phase I studies produced five monoclonal antibodies to Sta; these were used to demonstrate several ELISA test formats for rapidly measuring picogram to nanogram quantities of Sta. Phase II studies will aim to refine these assays in the microtiter plate and dispstick format. These assays will utilize various combinations of immobilized STA-BSA, and monoclonal antibody-enzyme conjugates. Considerable effort will be devoted to developing methods to allow for direct detection of STa in human and porcine stools. We will also begin preliminary work on an ELISA test for E. coli labile toxin (LT).The long-term Phase II goal is to develop a full panel of rapid and convenient diagnostic tests for enteric pathogens. Such tests are likely to become important new tools for basic and clinical research.National Institute of Allergy and Infectious Diseases (NIAID)