The in vitro excystation of Giardia lamblia on cysts isolated from human feces was studied. After purification by sucrose gradient, cysts were incubated in a pepsin-acid solution, then placed in a modified HSP3 medium where excystation occurred within a few minutes. The excystation procedure was studied by continuous observations by light microscopy and sequential observations by scanning electron microscopy (SEM). The in vitro excystation was stopped at timed intervals during incubation by addition of a large amount of 1% glutaraldehyde. The excystation process began by the cyst wall opening at one pole. Flagella protruded rapidly, the parasite emerged progressively from the cyst envelope, posterior end first, the empty cyst collapsed and shrank. Although flagella emerging from the organism were distinguishable, the cell body had not yet shown all the morphological features of the G. lamblia trophozoite. A radical rearrangement of the organism occurred gradually: initially oval in shape, the parasite became round, then elongated, flattened, and underwent cytokinesis. The daughter trophozoites acquired their typical morphological features: the shape, the adhesive disc with the C-shaped structure distinctly visible on the ventral surface, and the definite placement of the flagella. These observations obtained on G. lamblia by SEM were comparable to those obtained with G. muris.