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To better understand and study the infection of the protozoan parasite Cryptosporidium parvum, a more sensitive in vitro assay is required. In vivo, this parasite infects the epithelial cells of the microvilli layer in the small intestine. While cell infection models using colon, kidney, and stomach cells have been studied to understand the infectivity potential of the oocysts, an ideal in vitro model would be readily-available, human-derived, and originating from the small intestine. In this study, we developed a reproducible, quantitative infection model using a non-carcinoma, human small intestinal epithelial cell type, named FHs 74 Int. Our results show that FHs 74 Int cells are productively infected by viable oocysts, and exhibit higher levels of infection susceptibility compared to other cell types. Moreover, infection rate of the sporozoites on the monolayer was found to be comparable or better than other cell types. We furthermore demonstrate that infection can be improved by 65% when pre-treated oocysts are directly inoculated on cells, compared to inoculation of excysted sporozoites on cells. Identification of a better infection model, which captures the preferred site of infection in humans, will facilitate studies on the host pathogenesis mechanisms of this important parasitic human pathogen. Published by Elsevier B.V.