It has been reported that the numbers of people suffering from occupational asthma and skin rashes triggered by various chemicals in indoor air have increased markedly. Two-ethyl-hexanol (2-EH) is known to be an indoor air pollutant and its influence on health is of great concern. However, there are only a few reports regarding its effect on immune cells. Thus, we investigated the effects of 2-EH on immune responses in vitro with respect to effects on regulation of transcription factors as well as on 2-EH induced proliferation of spleen cells in vitro. The production of interleukin (IL)-6 and immunoglobulin were not induced by 2-EH. To characterize the effector cells of 2-EH, we prepared CD4-positive, CD8-positive, and peritoneal exudate cells (PEC). IL-2 was induced by 2-EH in CD4 cells, but not in CD8 cells. CD3-induced IL-2 expression was enhanced by 2-EH in CD4 cells, but not in CD8 cells. Moreover, IL-6 production was not induced by 2-EH in PEC. Nuclear factor-kappa B, nuclear factor of activated T, and signal transducer and activator of transcription-5 were activated by 2-EH in CD4 cells. Taken together, 2-EH activated CD4 cells, where this was accompanied by the activation of transcription factors. This suggested that the indoor pollutant 2-EH could function as a modulator of immune response.