Enteropathogenic Escherichia coli (EPEC) is a major aetiological agent of childhood diarrhoea in developing countries. The structural repeating protein A subunit, BfpA, found in the bundle-forming pilus, is one of the virulent factors for EPEC pathogenesis. Recombinant BfpA in laying hens elicited sustained and vigorous antibody production. Immunoglobulin Y (IgY) anti-BfpA antibodies were recovered from egg yolk, purified and characterized. Immunoadsorption with whole extracts of the isogenic E. coli EPEC adherence factor (EAF) strain that lacks BfpA rendered the resulting IgY preparations capable of: (a) recognizing purified or recombinant BfpA proteins in a dose-dependent fashion; (b) blocking the colonization of HeLa cells by EPEC EAF+, in vitro; (c) specifically identifying E. coli bearing EAF+; and (d) inhibiting the growth of E. coli EAF+ but not the EAF strain. IgY anti-BfpA is potentially useful as a specific, low-cost immunobiological reagent to screen human faecal specimens for the presence of EPEC.