Skip to main content
Campus Operations Banner

Autoclave 206

206. Use of Autoclave

A steam autoclave may be used to sterilize media, glassware, waste, instruments, etc. To accomplish the desired end goal and to protect the user and the environment from hazardous materials, the autoclave must be used correctly. Additionally, wastes must be managed in compliance with state and local regulations.

For the autoclave process to be effective in achieving sterilization, sufficient temperature, time and direct steam contact are essential. Air must be completely removed from the sterilizer chamber and from the materials to allow proper steam penetration. Factors that affect air removal include type and quantity of material to be autoclaved, packaging, load density and configuration, and container type, size, and shape.



  • All potentially infectious materials must be autoclaved before being washed and stored or disposed.
  • Personnel who use an autoclave must be trained to understand proper packaging, loading, labeling, and operation procedures.
  • Biohazardous materials must be labeled as such and must be sterilized by the end of each work day, or must be secured appropriately. Do not leave biohazardous materials in an autoclave overnight in anticipation of autoclaving the next day.
  • Do not autoclave materials that also contain toxic or volatile chemical or radiological agents.
  • Use bags or other containers labeled “Biohazard” for items that contain or may be contaminated with potentially infectious agents.
  • Use plain, unmarked containers for items that are not hazardous.
  • Do not double bag waste or tightly seal containers as this will impede steam penetration.
  • Do not put sharp objects such as broken glassware into an autoclave bag.
  • Open, shallow metal pans are more effective in conducting heat and allowing air removal than tall, plastic tubs.
  • Vessels with liquid should not be plugged or tightly capped.
  • It is advisable to add some water to bags of solid wastes (the water will vaporize into steam that will drive out residual air once sterilization temperature has been reached inside the bag).
  • When using an autoclave bag with a ‘Biohazard’ symbol on it, place a strip of tape that produces the word “autoclaved” across the symbol. This must be done for any autoclave bag that has a biohazard symbol.
  • Place containers of liquid, bags of agar plates, or other items to be autoclaved that may boil over or leak inside a secondary container of sufficient size.
  • Never place autoclave bags or glassware in direct contact with the bottom of the autoclave.
  • Do not overload the autoclave; leave sufficient room for thorough steam circulation.
  • Make sure the plug screen in the bottom of the autoclave is clean.
  • Do not mix loads of liquids with solids.
    1. The parameters for the sterilization cycle will depend upon the amount and type of material. Usually 121 °C at 15 psi for a minimum of 30 minutes is recommended. However, the temperature and cycle time can be increased taking into consideration the size and type of load. When in doubt, a biological indicator should be used as verification that sterilization was achieved (e.g. ampoule of Bacillus stearothermophilus spores placed in the middle of the full load). Document the date and results of any verification tests using biological indicators.
    2. If more than 50 pounds of waste per month is treated in the autoclave, efficacy MUST be monitored with a biological indicator at an appropriate frequency to ensure that the sterilization parameters are effective in treating biohazardous waste.
    3. Make sure chart paper or printer paper is in place to document the cycle parameters for the load. If a recording system is not available, it is critical to verify that sterilization parameters were achieved by another means such as spore strips, an autoclave thermometer, etc.
    4. The exact operating procedure for each model of autoclave will differ. The user should develop an SOP to describe proper steps to operate the autoclave.
  • After sterilization open the autoclave door no more than 0.5 inch and wait 10 minutes before unloading items.
  • Wear heat resistant gloves to unload items.
  • Be very careful of liquids, molten agar, etc. to avoid getting splashed with scalding liquid. Do not agitate containers of super-heated liquid or remove caps before unloading.
  • Unload hot items onto a cart for transport.
  • Attach a label to autoclaved waste that states, “Treated in accordance with the provisions of 25 TAC § 1.136(a)” and place the treated waste inside another opaque bag (black, green, etc.) to be placed in the regular trash.
  • NOTE: If a faulty condition exists (e.g. sterilizer did not finish the cycle, or water leaks out when the door is unlocked), contact a service technician.
  • Document the treatment of each load of biohazardous waste on the Autoclave Waste Treatment Record provided by EH&S which lists: the date of treatment, the amount of waste treated, the method/conditions of treatment, and the printed name and initials of the person performing the treatment. Keep charts or printout strips with the log sheets as documentation of the autoclave operation. These records will be inspected and collected by EH&S on a regular basis.
  • Put a strip of tape that produces the word “AUTOCLAVED” across all Biohazard symbols on bags. Do not throw away a biohazard bag without covering all biohazard symbols. 
  • Keep a log of each autoclave load of biohazardous waste. 
  • Replace chart paper or printer paper to record operating parameters of each autoclave load. 
  • When maintenance work or repairs are needed, the user must provide a safe work environment for the service technician. Remove all items from the sterilizer chamber, clean any spills or leaks inside the chamber, remove untreated biohazardous materials from the vicinity, etc.