In human chronic inflammatory bowel disease involving mucosal epithelium, sera and lamina propria mononuclear cells are reactive with cell surface components isolated from gut epithelial cells. To define a model system in which the disease-inducing potential of such immune factors could be rigorously evaluated, we sought to immunologically sensitize inbred murine strains to syngeneic colonic epithelial cell-associated components (ECAC-C), to define precise in vivo and in vitro conditions to optimize ECAC-C reactivity, and to initially explore whether such cells could elicit tissue injury in epithelium after adoptive transfer to naive animals. Following footpad immunization, Day 42 lymph node cells but not splenocytes were reactive with syngeneic ECAC-C, as shown by a linear increase in [3H]thymidine incorporation over a wide range of antigen concentration (0.5 to 100 micrograms/ml). A subsequent 48-hr exposure to ECAC-C and/or interleukin 2 resulted in a more restricted responsiveness, proliferation occurring only in the presence of ECAC-C or mitogen and not to a coimmunogen (PPD). Further evidence that lymph node cells from ECAC-C/CFA immunized animals were indeed sensitized to syngeneic ECAC-C included ability of donor animals to mount highly significant earlobe DTH responses to ECAC-C, indicating the presence of antigen-specific T-DTH cells, and the failure of polymyxin B, in doses sufficient to inhibit LPS-induced mitogenesis, to reduce lymph node cell responsiveness to ECAC-C, known to be contaminated with LPS. ECAC-C-specific circulating antibody and T-cytotoxic cells were not detected. Adoptive transfer of Day 42 lymph node cells, sensitized in vivo and conditioned in vitro, was not associated with tissue injury in syngeneic recipients in preliminary experiments. This model system, with antigen-specific cells analogous to those present in diseased mucosa of human chronic inflammatory bowel disease, may be an important means to determine the pathophysiologic significance of anti-epithelial cell immune responses in these disorders.