Objective To design an aerosol bacterial retention testing to evaluate bacterial retention capacity of air inlet filter of infusion device using challenging aerosol of Staphylococcus aureus. Methods Staphylococcus aureus suspension was diluted to 10⁸ CFU/mL, aerosol was produced by aerosol bacterial retention testing system, positive control branch and 75 air filters from 5 manufacturers (A-E) were tested, membrane filtration method was adopted to collect liquid specimen in the liquid impact sampler, bacteria growth and count was observed. Results After the testing, the collected fluid of positive control branch and 60 specimens from 12 batches of 5 manufacturers (A-D) were without bacteria growth and met the requirements; bacterial colonies of 15 specimens from 3 branches of ma-nufacturer E was >100 CFU, which did not meet the requirements. The air filter of manufacturer E was made of glass fiber with a pore diameter of 5 μm. Conclusion The designed aerosol bacterial retention testing system can successfully retain the bacteria in the air inlet filter of infusion devices.