Autoclave Procedure

Wear personal protective equipment:

  • Lab coat
  • Eye protection
  • Closed-toe shoes
  • Heat-resistant gloves to remove items, especially hot glassware

Packaging and Loading

  • Only designated individuals should be allowed to set and/or change parameters for the autoclaves.
  • Before using the autoclave, check inside for any items left by the previous user that could pose a hazard.
  • Clean the drain strainer before loading the autoclave.
  • Always place items in a secondary container.
  • Do not overload or package bags too tightly. Leave sufficient room for steam circulation. If necessary, place container on its side to maximize steam penetration and avoid entrapment of air.
  • Use only autoclavable bags to package waste.
  • Do not allow bags to touch the interior walls of the autoclave to avoid melting of plastic.
  • Ensure sufficient liquid is packed with contents of autoclave bags if dry.
  • Place soiled glassware and lab ware in secondary containers and autoclave them in the solids cycle. Do not fill containers more than 2/3 full with liquids. Loosen caps or use vented closures.
  • In case of clean glassware and wrapped instruments, lay them in a secondary container before autoclaving in wrapped goods cycle.
  • For secondary containment, use autoclave trays made out of polypropylene, polycarbonate or stainless steel. The trays should have a solid bottom and sides to contain the contents and catch spills.
  • Choose appropriate cycle for the material.  Incorrect selection of cycle may damage the autoclave, cause liquid to boil over or bottles to break.
  • Start your cycle and fill out the autoclave user log.  A completed cycle usually takes between 1 to 1.5 hours.
  • Check chamber/jacket pressure gauge for minimum pressure of 20 pounds per square inch (psi).
  • Close and lock door.
  • Check temperature for 250⁰F (121⁰C) every load.
  • Do not attempt to open the door while autoclave is operating.


  • Ensure cycle has completed and both temperature and pressure have returned to a safe range.
  • Wear PPE described above, plus an apron and face shield if removing liquids.  Stand back from the door as a precaution and carefully open door no more than 1 inch. This will release residual steam and allow pressure within liquids and containers to normalize.
  • Allow the autoclaved load to stand for 10 minutes in the chamber. This will allow steam to clear and trapped air to escape from hot liquids, reducing risk to operator.
  • Do not agitate containers of super-heated liquids or remove caps before unloading.
  • Place liquids in an area which clearly indicates the items are “hot” until the items cool to room temp.
  • Allow autoclaved materials to cool to room temperature before transporting. Never transport superheated materials.
  • Place cooled autoclaved biohazard bag into regulated medical waste box.  Autoclaved infectious liquids may be disposed of into the sanitary sewer.


Meagan Fitzpatrick
Biosafety Officer

Halina Staniszewska,
Sr. Lab Safety Specialist