A major constraint in the development of methods for the identification of chemical respiratory allergens is the continuing uncertainty regarding the mechanisms of this disease and in particular the role of IgE antibody. There is, however, increasing evidence that respiratory sensitization is favoured by the induction of a selective type 2 cytokine response. The current investigations focus on the potential application of cytokine profiling to the identification of chemical respiratory sensitizers. The objective is to determine the optimal configuration of the method for discrimination between chemical contact and respiratory sensitizers. The reference contact sensitizer 2,4-dinitrochlorobenzene (DNCB) and reference respiratory sensitizer trimellitic anhydride (TMA), which have been shown to induce type 1 and type 2 cytokine profiles, respectively, were utilized. Variables investigated included cell concentration, time in culture, dosing regimens (a 13 day and a truncated 8 day protocol) and the impact of restimulation in vitro with T cell mitogens. Cell culture conditions were critical for the selectivity of the response, with the addition of mitogen resulting in a less discriminatory pattern of cytokine expression, particularly for the type 1 cytokine interferon gamma (IFN-gamma). Furthermore, a 13 day exposure period was required for vigorous expression of IFN-gamma by DNCB-activated cells, whereas type 2 cytokine expression by TMA-stimulated cells was recorded after 8 days. These data demonstrate that the most optimal method for cytokine profiling is a chronic (13 day) exposure regime followed by culture of lymph node cells at 10(7 )cells ml(-1) for 120 h in the absence of mitogen.