Pancreatic digests of casein are major ingredients of media used in the axenic cultivation of lumen-dwelling parasitic protozoa, especially Entamoeba, Giardia, and trichomonads. The digest used almost exclusively in the development of these media, Medo-Peptone (Trypticase BBL), has not been available since 1981. Moreover, none of dozens of similar type digests tested since then in our laboratory has proved equal to Medo-Peptone, and in the last two years it has become increasingly difficult to obtain new batches which will support even modest growth of Entamoeba histolytica. In response to this problem we have developed a casein-free medium, YI-S, consisting of a nutrient broth, vitamin mixture and serum. We recommend it as a replacement for the casein-dependent medium TYI-S-33, currently the most widely used for axenic culture of Entamoeba histolytica and other lumen-dwellers.