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Safe Use of Autoclave Sterilizers

Autoclaves use heat and pressure with water to create superheated steam. Therefore, they can pose significant hazards to untrained or careless employees

Vince McLeod, CIH

Vince McLeod is an American Board of Industrial Hygiene-certified industrial hygienist and the senior industrial hygienist with Ascend Environmental + Health Hygiene LLC in Winter Garden, Florida. He has more...

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Autoclaves are so common and familiar in labs today that it is easy to overlook the associated hazards. Consider these recent incidents reported by the Lab Health & Safety Committee of the American Industrial Hygiene Association.1

A sudden explosion rocked a laboratory building one morning. A search by lab personnel found that an autoclave had ruptured, blowing its 80-lb door off its hinges, across the room, and into the opposite wall. Luckily, no one was in the room when the door blew off. With racks of test tubes, stacks of culture media, and trays of equipment (including needles) awaiting sterilization prior to disposal splattered across the room, it looked as if a bomb had gone off. Further investigation was unable to determine the cause of the failure. Not very reassuring, to say the least.

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In another incident, a post-doc was severely scalded while removing a load from a tower-style autoclave. She was attempting to remove a Nalgene tub containing oneliter bottles of media in water when the tub buckled and spilled near-boiling water onto her torso and thighs. She was not aware of the contents of the Nalgene tub because the autoclave log had not been filled out for the load.

And in a third incident, two steam releases occurred in an autoclave room. One was due to a lack of a backflow preventer, which allowed steam to backflow into a DI water feed made of PVC. The PVC melted, releasing boiling-hot steam into the room. The second release occurred in the same room when the automatic overpressure safety valve switch failed. It was found that the electrical safety switch had been damaged during the previous release.

It is easy to get hurt when you are dealing with pressurized steam, but you can avoid mishaps and potential significant damage or injury by following a few basic safety precautions.

Recognizing the hazards

As a start, always think of autoclaves as large specialized pressure cookers. Do you remember all the warnings and stories your mother and grandmother told you about using pressure cookers? If not, consider that autoclaves use heat and pressure with water to create superheated steam. Therefore, they can pose significant hazards to untrained or careless employees.

Autoclaves are basically used for two main purposes—either to steam-sterilize media, instruments, or lab equipment such as glassware and specialized implements or to inactivate biological waste materials.2 As we have indicated, the primary hazards are physical ones presented by high temperatures, steam, and pressure. Effective sterilization requires steam temperatures in excess of 250°F (121°C). Typical autoclave pressurization is at least 20 pounds per square inch. Depending on the use, additional concerns may include biological hazards such as infectious materials or physical hazards from sharps.

First—train, train, train

Loading and running an autoclave may seem as simple as using your dishwasher at home, but some planning is required to operate the autoclave safely and efficiently. All operators should be thoroughly familiar with the owner/operator’s manual and controls. Controls vary between manufacturers, and every machine has unique loading characteristics, load-sizing requirements, cycle settings, and cycle types.3 The size of the load and types of materials requiring sterilization or inactivation/decontamination will determine the cycle needed. It is highly recommended that the manufacturer’s operation manual be copied and waterproofed/ laminated and kept in the room with the autoclave.

Adopt a policy in which all users must be trained prior to operating any autoclave. Principal investigators or laboratory supervisors must bear the responsibility of ensuring this is done. Ensure all training is documented, and maintain the records in the lab. Make sure your training addresses proper use of appropriate personal protective equipment.

At a minimum, training should cover:

  • Location, function, and use of controls
  • Proper loading and unloading (including packaging, sizing, and testing protocols)
  • Required personal protective equipment (heat-resistant gloves, lab coats, eye protection, and closed-toe shoes)
  • Incident and maintenance reporting and record keeping
  • Emergency procedures

Second—develop and enforce a monitoring and testing protocol

To ensure the autoclave is functioning properly and sterilization/ inactivation is effective, we should monitor the operation of the autoclave and routinely test sterilization cycles. In fact, in Florida, this is mandated by the Florida Administrative Code for handling biomedical wastes, FAC 64E-1.4 Under this law, autoclaves must be tested before being placed into service and routinely afterward. For autoclaves used to inactivate substances such as human pathogens, blood, tissues, and clinical samples, testing is required after every 40 hours of use. Autoclaves used to sterilize other materials must be tested every six months. We believe this is a reasonable testing schedule for research laboratories. Other institutions recommend testing with biological indicators at least once per month.3

Testing an autoclave’s sterilization effectiveness requires the use of biological indicators. These are available in commercially prepared test kits containing bacterial spores, usually Bacillus stearothermophilus (e.g., ProSpore2). Most spore vial test kits require incubation of the autoclaved test vial along with a nonautoclaved control vial. Incubation will allow surviving spores to grow. We recommend that test loads, if used, approximate the weight and density of actual waste or materials normally autoclaved. For best results, test vials should be placed at the bottom, top, front, rear, and center of the autoclave chamber by placing vials in those positions of the test load or by making a number of smaller test packs with vials in the center and placing the packs appropriately in the chamber. In this way, the correct parameters for sterilization (time, temperature, and pressure) can be determined.

Third and final step—record keeping

A good autoclave safety program must include documentation. Principal investigators and supervisors are responsible for ensuring proper records are kept up to date. Autoclave users should be responsible for recording autoclave run information.

In addition, we recommend keeping records of all on-site maintenance. Only contractors approved by the manufacturer should perform maintenance. Keep maintenance contractor contact information conveniently posted.

Log each load processed in the autoclave. Record the date, time, and operator’s name and contact information (e.g., lab, room number, and phone number). Indicate whether the load is biohazardous material and record the temperature, pressure, and time length for the cycle. If the autoclave provides printed data or if data is recorded on a cycle wheel, save the printout or disk. Finally, include information in the log sheet for all efficiency tests performed and the results of each test.

Summing up

Additional information and technical assistance are always available from manufacturers as well as from NIOSH, OSHA, and many academic websites. The key to working with autoclaves is first recognizing the hazards, followed by training, testing, and record keeping. Whenever possible, we recommend autoclaves be located in dedicated rooms with proper and sufficient facility supply and exhaust ventilation.


1. Lab Safety Autoclaves Incidents, Lab Health & Safety Committee, American Industrial Hygiene Association. Falls Church, VA. 2018. 

2. Safe and Effective Use of Autoclaves. Environmental, Health and Safety Fact Sheet #33. University of California at Berkeley. 2011. 

3. Environmental Health and Safety Update – Autoclave Safety. Environmental Health and Safety. Weill Cornell Medical College. 2003. 

4. Biomedical Waste Program, Florida Department of Health, Bureau of Community Environmental Health. 2002.