Steam Sterilizer (Autoclave) Use

The steam sterilizer (autoclave) is a steam-pressure vessel, which must be operated in a proper manner to ensure inactivation of microbial agents and the safety of the operator. A steam autoclave is a gravity flow sterilizer, which must reach a minimum temperature of 121°C (250°F) at 15 pounds per square inch (psi) of internal steam pressure. Depending on the volume and density, the biohazardous material may require 60-120 minutes for sterilization.

DSOs/LSOs must provide training to research personnel on the correct preparation of biohazardous materials for sterilization, and on the correct use of steam autoclaves. Autoclave users should understand the time, temperature, pressure relationships required for proper decontamination of biohazardous materials. Users should complete the CITI training course, Autoclave Safety Training, prior to using an autoclave.

Personnel should wear proper personal protective equipment when operating the autoclave, particularly when unloading the autoclave, including:

  • Lab coat
  • Heat resistant gloves
  • Closed-toe shoes
  • Eye protection

Steam autoclave sterilizers should be certified during annual preventative maintenance. Autoclave door clamps and seals should be inspected for wear and damage over time. Prior to using a steam autoclave, inspect the autoclave components for proper operation and remove any debris from the strainer for the autoclave chamber floor drain.

After the steam autoclave completes a sterilization cycle, check the printout to confirm the cycle reached the correct temperature and pressure for the configured exposure time. If the printout indicates an aborted autoclave cycle, then assume that the contents are still potentially biohazardous.

Steam Sterilization Process Validation

It really is important when autoclaving waste to have validated the process with biological indicators included inside surrogate loads that represent normal biohazardous waste (i.e. number, content, and position of autoclave-safe biohazard bags). Biological indicator systems based on Geobacillus stearothermophilus spores are designed to demonstrate that the programmed steam autoclave cycle effectively inactivates microorganisms. These systems assure that the right combination of Time, Temperature, and Steam Saturation occurred during the cycle to ensure sterilization.

Steam Sterilization Process Controls

Heat-sensitive autoclave tape is a type of chemical indicator that only confirms that the steam sterilizer reached the minimum processing temperature. A change in color of heat-sensitive tape does not indicate that the steam autoclave cycle has decontaminated the biohazardous materials.

The Biosafety Program recommends the use of 3M™ Comply™ Thermalog™ or 3M™ Comply™ SteriGage™ chemical integrators to confirm the sterilization process. Chemical integrators provide immediate, easy to read “Accept” or “Reject” sterilization results. You can purchase these chemical integrators from any major commercial laboratory supply vendor.

EPA-Recommended Decontamination Processing (Dwell) Time

Dwell Time

Trash (paper toweling, etc.)

60 minutes


60 minutes


60 minutes/4 Liters

Animal Carcasses

120 minutes**

Animal Bedding

120 minutes

** The US EPA does not accept autoclaving as a terminal sterilization for animal carcasses. The process renders the carcasses safe for collection in fiber drums for terminal incineration off-site.