SUMMARY: Teehniques are deseribed for the dissociation, fractionation through Percoll, and in vitro maintenance of mucosal epithelia from the human or guinea pig large bowel. Tissue recovered after surgeu is predigested with trypsin-citrate and treated subsequently with a mixturc of trypsin, citrate, and collagenase. The resulting suspension of single cells, cell clusters, and partially digested crypts is suspended over a Percoll solution and enriched in multicellular elements by two sequential centrifugations. The recovered multicellular complexes are inoculated to specially treated culture vessels in a serum-free medium supplemented with epidermal growth factor, insulin, transferrin, selenium, and bovine pituitary extract. Epithelia, characterized as such by transmission electron microscopy, adhere to the substrate, form colonies, and can be maintained routinely for study for at least 10 wk.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.