NEW APPROACHES TO THE DIAGNOSIS OF AMEBIASIS

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Amebiasis, caused by Entamoeba histolytica, is a major problem in developing countries. In developed countries, it is most common in immigrants and travelers returning from the third world, and less common but present in men who have sex with men and immunosuppressed individuals. In previously sponsored SBIR research, we developed a stool antigen test for the detection of the E. histolytica adhesin, a protein that is an effective in vitro diagnostic marker. Our test, a microwell ELISA, is more accurate than microscopy and other commercial ELISAs, and more rapid than the "gold standard" culture/zymodeme, which takes 2 weeks to complete. We now have developed antigen affinity-purified antibody that has enabled us to increase the sensitivity' of the test ten-fold. We propose to use the improved antibody in dipsticks that can be used in the field as a rapid stool antigen test. In an effort to make immunoassays available to laboratories that test only fixed specimens (where most misdiagnosis of amebiasis occurs in the U.S.), we will develop immunoassays that detect formalin-stable Entamoeba antigens. We also propose to develop an antibody-based test for Entamoeba cysts. Polyclonal and monoclonal antibodies against cysts will be developed and used in screening immunoassays for contaminated water and food (recent food and water- borne outbreaks in Reston Virginia and Tblissi, Republic of Georgia respectively, make this a topical and immediate need). The development of cyst-specific tests will be patterned after those for the cyst- and oocyst- specific antigens of Giardia and Cryptosporidium. The data collected will be used to develop 510(k) documents for submission to and review by the FDA. This project will lead to a battery of tests that have broad application in the diagnosis of amebiasis. The tests will be crucial tools for evaluation of amebiasis vaccine candidates developed under our separate Tropical Disease Research Unit. PROPOSED COMMERCIAL APPLICATIONS: New diagnostic tests will be developed for amebiasis to solve the problems of misdiagnosis at community and reference labs in the U.S., enable screening of water supplies for the parasite, and to permit field evaluation of vaccine trials. Our results should lead to commercial tests that will be of significant value to the health industry around the world.