Aims The aim of this study was to demonstrate a prototype tool for measuring infectivity of an aerosolized human pathogen influenza A/PR/8/34 (H1N1) virus using a small-animal model in the Controlled Aerosol Test System (CATS). Methods and Results Intranasal inoculation of nonadapted H1N1 virus into C57BL, BALB/c and CD-1 mice caused infection in all three species. Respiratory exposure of CD-1 mice to the aerosolized virus at graduated doses was accomplished in a modified rodent exposure apparatus. Weight change was recorded for 7 days postexposure, and viral populations in lung tissue homogenates were measured post mortem by DNA amplification (qRT-PCR), direct fluorescence and microscopic evaluation of cytopathic effect. Plots of weight change and of PCR cycle threshold vs delivered dose were linear to threshold doses of 40 TCID50 and 12 TCID50, respectively. Conclusions MID50 for inspired H1N1 aerosols in CD-1 mice is between 12 and 40 TCID50; proportionality to dose of weight loss and viral populations makes the CD-1 mouse a useful model for measuring infectivity by inhalation. Significance and Impact of the Study In the CATS, this mousevirus model provides the first quantitative method to evaluate the ability of respiratory protective technologies to attenuate the infectivity of an inspired pathogenic aerosol.