Clonal Growth of Entamoeba histolytica and other Species of Entamoeba in Agar*

This is the first description of a method for growing axenized Entamoeba histolytica as colonies from single cells in agar suspension. Factors affecting the efficiency of colony formation included unknown characteristics of the amebal strain, age of cells used, and the type and concentration of agar employed. Colony formation was dependent upon filling deep vessels (culture tubes or tissue culture flasks) with agar, probably to maintain reduced oxygen tension, and upon solidifying the agar rapidly at 0 C. It was demonstrated in a study with the drug metronidazole that the agar colony method was useful for quantifying cell viability in drug testing. Eleven of 12 E. histolytica strains tested, as well as one Entamoeba terrapinae, one Entamoeba barreti, and 3 Entamoeba invadens strains formed colonies in agar suspension. E. histolytica-like amebae (Laredo type) and E. moshkovskii formed only tiny colonies in agar.