BackgroundUsing a rotary instrument or ultrasonic instrument for tooth preparation is a basic operation in the dental clinic that can produce a significant number of droplets and aerosols. The dental droplet and aerosol can lead to the transfer of harmful germs. The goal of this study was to analyze the properties of microbiological aerosol created by droplets and aerosol generated by three common tooth-preparation instruments.MethodsStreptococcus mutansUA159 was used as the biological tracer to visualize the droplets and aerosols. The passive sampling method was used to map the three-dimensional spatial distribution and the six-stage Andersen microbial sampler (AMS) was used as the active sampling method to catch aerosol particles at a specific time.ResultsThe aerosol concentration is related to instruments, three-dimensional spatial distribution, and dissipation time. Most aerosols were generated by air turbines. More microorganisms are concentrated at the 1.5 m plane. The majority of the post dental procedure contamination was detected within the 0–10-min period and it decreased rapidly within 30 min.ConclusionThis study is conducive to the proposal and improvement of relevant infection control measures in dental procedures and provides a basis for the assessment of measures, reducing the risk of nosocomial infection.