Isolation and purification of Giardia trophozoites from rat intestine

A method for the isolation of Giardia trophozoites based on their ability to attach to warm surfaces has been developed. Mucosal scrapings were obtained from the small intestine of rats infected with Giardia and suspended in Hanks' balanced salt solution (HBSS). Trophozoites were concentrated by centrifugation and allowed to attach to the surfaces of polystyrene petri dishes incubated at 37 C. Incubation temperature significantly affected the recovery of trophozoites. After attachment at 37 C, trophozoites were separated from contaminating intestinal debris by incubation at cold temperature. The trophozoites detach at 4 C, whereas the intestinal debris remain adherent. Then the detached trophozoites were isolated by reattachment at 37 C. Examination by scanning electron microscopy revealed a marked reduction in contamination of attached trophozoites and dish surfaces after the use of cold temperature detachment and reattachment at 37 C. Viability of trophozoites as measured by erythrosin-B dye exclusion, remained above 90% up to 120 min after isolation. This method of isolation facilitates the recovery of this protozoan directly from small intestine for morphological and experimental study.