The effects of ambient levels of NO2on the spleens of adult and newborn Swiss Webster (S/W) mice were determined. Spleens were evaluated by the following criteria: (1) spleen weight, expressed as a percent of the body weight (% spleen weight), (2) size of the spleen lymphoid nodules as determined by computed image analysis, (3) spleen cell counts, and (4) histopathologic evaluation. Data for NO2‐exposed animals were compared with data for control animals that inhaled filtered air. Totals of 217 control and 217 NO2‐exposed animals were studied. After 6 wk of exposure to NO2at ambient levels (0.35 ± 0.05 ppm), the following significant changes were observed in the spleens of exposed mice: (1) increase in % spleen weight (p <0.0025), (2) increase in size of spleen lymphoid nodules (p <0.01), (3) smaller increase in spleen cell number per given weight increment of spleen as determined by correlation coefficients (p < 0.0125) and linear regression analysis of spleen cell counts, and (4) an apparently greater predominance of red cells in the red pulp. It is concluded that inhalation of N02is associated with quantifiable spleen changes, which may prove to be useful indicators for assessing effects of inhaled NO2.