Multiplex Immunodiagnostic Panel for Category B Agents of Diarrhea and Dysentery
Our goal is to produce a multiplex immunodiagnostic panel specific for Cryptosporidium, Giardia, E. histolytica, and shiga toxin-producing Shigella and E. coli detection in stool. Additional effort will quantify the fecal biomarkers lactoferrin and hemoglobin, indicators of intestinal inflammation and hemorrhage. The full multiplex panel will provide rapid diagnosis of major Category B enteropathogenic agents of diarrhea/dysentery and describe severity of disease.
The assay utilizes sensitive and specific immunofluorescence-based Luminex technology. The Luminex allows multiplexed detection of analytes in a single sample using an array of uniquely detectable bead sets. Analyte-specific antibodies are selected from existing FDA-cleared immunoassays, providing assurance of their performance and stability. Preliminary data indicate Luminex sensitivity for the enteric protozoa exceeds that of ELISA, and is comparable to RT-PCR, with purified parasite curves. Detection in stool identified 90 of 90 parasite ELISA-positive reactions, but still requires complete validation of clinical sensitivity. Luminex detection of lactoferrin and hemoglobin correlate with commercial ELISA detection and provide an expanded range of quantification, increasing efficiency of fecal analysis over ELISA.
1. Aim #1 details an enteric protozoan parasite panel for Cryptosporidium, Giardia, and E. histolytica.
2. Aim #2 expands this diarrhea/dysentery panel to include detection of shiga toxin, shiga-like toxin 1 (Stx1) and shiga-like toxin 2 (Stx2).
3. Aim #3 adds detection of lactoferrin and hemoglobin as biomarkers of diarrhea/dysentery severity.
Successful completion of these studies will provide a prototype diagnostic capable of the simultaneous detection of Category B enteropathogens and intestinal biomarkers that describe severity of infection. The multiplex assay will be competitive for NIH small business funding to complete FDA 510(k) clearance validations for commercial marketing. The Luminex-based multiplex detection is an excellent candidate for commercialization, and it allows for future inclusion of other enteric pathogens and gut function biomarkers.
Joel Herbein