An antigen-capture enzyme-linked immunosorbent assay employing rabbit and mouse antisera to Giardia lamblia cyst antigens was developed for the diagnosis of Giardia infection through detection of G. lamblia-specific stool antigens in cell-free aqueous eluates of human stool. This is the first report of the use of anti-cyst antibodies in an enzyme immunoassay for G. lamblia. The assay gave a positive result with 54 of 59 stools from patients with symptomatic, clinically diagnosed giardiasis, giving the test a sensitivity of 91.5%. A negative reading was obtained with all of 25 stools from G. lamblia-negative control patients. The assay could detect as few as 20 sonicated cysts added to control stool eluate. The assay was more sensitive to cyst-derived antigens than to trophozoite-derived antigens. With two exceptions, the assay gave a negative result with stools from patients infected with Entamoeba histolytica (seven), Cryptosporidium sp. (four), or Blastocystis hominis (seven) and thus appears to be specific for G. lamblia antigens. Storage of stool eluates for more than 6 months at 4 degrees C as unpreserved aqueous eluates or as formalinized eluates did not affect the ability of the assay to detect the giardial antigens. The enzyme-linked immunosorbent assay proved useful for monitoring the levels of G. lamblia-specific stool antigens in the stool of patients undergoing antigiardial chemotherapy.