It has been shown that pulmonary exposure to diesel exhaust particles (DEP) disrupt immune systems, presenting as exacerbating effects on allergic manifestations (i.e., allergic asthma). However, since inhalation system could not be developed, impact of nano-level DEP on health has not been satisfactorily elucidated. Our institute (National Institute for Environmental Studies) established the "environmental nanoparticle exposure system applied in animals" in 2005, and, since then, we have explored the health effects of the exposure. As part of our ongoing research, the present study was aimed to investigate the effects of nanoparticle-rich DEP (NRDEP) on the characterization of primary atopy-prone splenocytes in vitro. NC/Nga mouse-derived splenic mononuclear cells were co-cultured with NRDEP (0-50 µg/ml); thereafter, the surface expression of CD11c, CD80, CD86, CD69, and CD40L was evaluated by means of flow cytometry. NRDEP increased the surface expression of these molecules on the splenocytes in a dose-dependent manner with an overall trend (with significance vs. 50 µg/ml of NRDEP). These results suggest that NRDEP can activate naïve splenic mononuclear cells from atopy-prone animals.