Abstract
An intestinal protozoan parasite, Cryptosporidium parvum, is a major cause of waterborne gastrointestinal disease worldwide. Detection of Cryptosporidium oocysts in potable water is a high priority for the water treatment industry to reduce potential outbreaks among the consumer populace. Anti-Cryptosporidium oocyst polyclonal and monoclonal antibodies were tested as capture and detection reagents for use in a fiber optic biosensor assay for the detection of Cryptosporidium oocysts. Antibodies were validated using enzyme-linked immunosorbent assays, flow cytometry, Western blotting and fluorescent microscopy. Oocysts could be detected at a concentration of 105 oocysts/ml when the polyclonal antibodies were used as the capture and detection reagents. When oocysts were boiled prior to detection, a ten-fold increase in sensitivity was achieved using the polyclonal antibody. Western blotting and immunofluorescence revealed that both the monoclonal and polyclonal antibodies recognize a large (>300 kDa) molecular weight mucin-like antigen present on the surface of the oocyst wall. The polyclonal antibody also reacted with a small (105 kDa) molecular weight antigen that was present in boiled samples of oocysts. Preliminary steps to design an in-line biosensor assay system have shown that oocysts would have to be concentrated from water samples and heat treated to allow detection by a biosensor assay.
Highlights
Cryptosporidium parvum is an intestinal protozoan parasite that continues to be a major cause of waterborne gastrointestinal disease worldwide
Effect of oocyst pre-treatment on antibody binding The anti-Cryptosporidium polyclonal antibody was first evaluated for detection of Cryptosporidium oocysts using enzyme-linked immunosorbent assays (ELISAs)
Many articles have been written that focus on various methods for detecting oocysts in water, yet most if not all deal with methods that rely upon grab samples, without the potential for advancing to in-line detection
Summary
Cryptosporidium parvum is an intestinal protozoan parasite that continues to be a major cause of waterborne gastrointestinal disease worldwide. Detection and removal of Cryptosporidium oocysts in potable water are high priorities for the water treatment industry to reduce potential outbreaks among the consumer populace. In the widely publicized 1993 Milwaukee Cryptosporidium outbreak, oocysts passed through the filtration system of one of the city's water treatment plants and an estimated 403,000 people suffered from gastroenteritis [1]. The breakdown of the filtration process was found to be related to high turbidity values of the water. The United States Environmental Protection Agency (EPA) regulatory approaches for Cryptosporidium removal in drinking water are currently based on filtration with compliance met using turbidity standards. Detection of Cryptosporidium oocysts in water using conventional microbial analysis is (page number not for citation purposes)
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