This study was designed to determine the effectiveness of the nebulisers’ interactions with SARS-CoV-2, to understand and inform the design of effective animal models utilising an aerosol infection route. Determination of spray factor allows one to calculate the presented dose to an animal, based on any given concentration of a pathogen in a nebuliser and is specific both to the aerosol challenge system (apparatus, mode of aerosolization, etc.) and the individual organism. For example, another respiratory pathogen, Influenza H1N1, delivered via a 6-jet Collison in the same Biaera/Hendersen system as used here has a SF of 1.32 × 10−6 [39,40] which is comparable to our derived value for SARS-CoV-2 of 2.23 × 10−6 in the same system. Our study also showed that the SLAG, mesh and jet nebulisers were no more likely to inactivate SARS-CoV-2 than either of the more standard Collison nebulisers, within our system. The lack of significant influence of RH, or even nebuliser type (excluding the 1-inch SLAG) on spray factors, demonstrates that the practice of using Collison nebulisers is predicted to not have a negative impact on the viability of particles generated in a SARS-CoV-2 infection model. Specific spray factor values calculated in this study will be used to determine the starting inoculum in an aerosol challenge system, required to give a specific presented dose (or range of doses) to animals. These data will allow improved and reproducible infection and interpretation of disease development and assessment of interventions in this model.