Giardia lamblia trophozoites specifically colonize the upper human small intestine which is normally serum-free, but grow in vitro only in medium supplemented with serum or serum fractions. Recently, biliary lipids were shown to support the growth of G. lamblia without serum. Now, it is reported that human duodenaljejunal mucus stimulates growth of Giardia in medium with biliary lipids. Stimulation by mucus was enhanced by inclusion of chymotrypsin or crude pancreatic proteases. Coculture of trophozoites with human intestinal epithelial cells also promoted growth, especially in the presence of mucus and/or biliary lipids. With biliary lipids alone, the mean increase in cell number was 3.2 fold and in the presence of mucus 8 fold (p < 0.01) in 24 serial subcultures. The demonstration that human intestinal mucus and epithelial cells promote serum-free growth of G. lamblia may help to explain specific colonization of the small intestine by G. lamblia.