Objective To investigate whether the animal nose?only inhalation exposure device can operate normally and produce aerosol particles of a specific size, to explore the air tightness and uniform distribution of the aerosol during operation, and to examine the disinfection effect of disinfectant on the system. Methods Escherichia coli (E. coli) aerosol and mouse hepatitis virus MHV?JHM aerosol were developed by the nose?only inhalation exposure device, and a solution of 75% ethanol and 3% hydrogen peroxide was run through the nebulizer to disinfect the system. E. coli and MHV?JHM were collected respectively in the exposure chamber, at sites where leaking easily occurred, including after disinfection. The sampled E. coli was cultured. In addition, RNA from the sampled MHV?JHM was extracted and one?step RT?PCR was then used to test the presence of viral RNA from the samples. Results The result showed that the mass median aerodynamic diameter of the E. coli aerosol was 1. 27 ± 0. 61 μm, that colonies grew after coating of a plate with sample liquid collected in the exposure chamber, that the numbers of cultured colonies by NC membrane sampling of different ports were similar, and that no colony growth was found on the sedimentation plate at sites where leaking readily occurred or on the sampling plates after disinfection. There was a positive result for MHV?JHM virus RNA of the sampling solution from the liquid impinger, while there were negative results for MHV?JHM virus RNA of the sampling solutions from the sites where leaking easily occurred and after disinfection. Conclusions The results show that the animal nose?only inhalation exposure device can operate normally and develop an aerosol that is passed through and distributed evenly in each port of the exposure chamber during operation; moreover, the device was shown to have good air tightness. The system can be effectively disinfected by using the corresponding disinfectant, and the disinfection effect is comprehensive.