Initial epidemiologic studies of acquired immunodeficiency syndrome (AIDS) occurring in homosexual men identified the use of the inhalants amyl, butyl, and isobutyl nitrite as possible risk factors contributing to the disease. Because of the lack of immunotoxicological data on these chemicals, we studied the effects of subchronic exposure to isobutyl nitrite (IBN) on the immune system. BALB/c mice were exposed to either 50 or 300 ppm IBN for 6.5 h/d, 5 d/wk for up to 18 wk. After 7, 13, or 18 wk of exposure, mice were killed and the following assays were performed. Antibody producing cells were enumerated by a slide plaque assay on animals immunized with sheep red blood cells while still in exposure chambers. The lymphocyte proliferative response to mitogens (phytohemagglutinin, concanavalin A, pokeweed mitogen, and lipopolysaccharide) was tested using several concentrations of each mitogen. Additional mice were immunized with Freund's complete adjuvant 21 d prior to death and were tested for delayed hypersensitivity response to purified protein derivative by a radiometric skin test. Finally, the relative numbers of T cells and T-cell subsets among splenic lymphocytes from exposed and control animals were determined. At the time periods tested there were no discernable immunotoxic effects observed in the exposed animals in any of the assays performed. These results indicate that IBN, at the dosages tested, had no discernable detrimental effect on the immune system of mice.