This study was designed to elucidate the mechanism of the leukocyte adherence inhibition (LAI) test in man. To identify the reactive cell types, enriched leukocyte populations (dextran-separated leukocytes and Hypaque-Ficoll-isolated mononuclear cells and neutrophils, as well as rosette-isolated B- and T-lymphocytes) were tested for leukocyte adherence in the absence of serum to tumor-specific antigens. LAI reactivity was not restricted to any of the enriched populations, suggesting the involvement of multiple cell types. Attempts to demonstrate soluble lymphocyte factors in the LAI mechanism have been uniformly negative. In contrast, factors in serum of immune donors were able to arm naive cells to be specifically responsive. This suggests a role for serum factors in the mechanism of LAI reactivity and partially explains the participation of multiple cell types in the responses observed. In additional studies, we could not document a correlation between the magnitude of the dermal test (delayed cutaneous hypersensitivity) and the magnitude of the LAI response in patients with squamous cell carcinoma of the head and neck. In 34 of 54 of these patients, there was agreement between the two tests (both positive, 27 of 54; both negative, 7 of 54). In the remaining 20 patients, the dermal test was greater than 5 mm while the LAI test was negative (less than 30% inhibition).