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Autoclave Safety and Use

Autoclave Training

Before using an autoclave, you must be trained by the Chemical Safety Coordinator.  To request training, please email Dr Moore at moore.4061@osu.edu.

Some Potential Autoclave Hazards 

  • Broken glassware
  • Super-heated liquids
  • Heat burns
  • Steam Burns
  • Scalds from hot liquid
  • Explosions

Secondary Containment

You are required to use secondary containment each time that you use an autoclave.

Polypropylene or stainless steel tubs are normally used for secondary containment.

Make sure your Plastic Container is Suitable for Autoclaving

You can use the following plastics:

  • Polypropylene (PP)
  • Polycarbonate (PC)

Do not use:

  • Polyethylene (PE)
  • High Density Polyethylene (HDPE)

 Other Secondary Containment Considerations

Use a container with the lowest side and widest diameter.

Leave space between items to allow steam to circulate.

Do not overload the chamber.

PPE 

Always wear proper PPE when using the autoclave

  • Lab coat
  • Heat resistant gloves
  • Eye protection
  • Face shield  (splash)
  • Liquid resistant apron

Before Using the Autoclave

  • Check Gaskets and seals to ensure they are undamaged and free of debris or dirt
  • Check inside for any leftover items
  • Check the drain strainer- for efficient heat transfer steam must flush air out of autoclave chamber.  If the drain screen is blocked , a layer of air may form and prevent proper autoclave operation

Preparing items for Sterilization 

  • Glassware
    • Cap Pyrex bottles loosely – whether empty or filled – to prevent explosions due to expansion.
    • Cover bottles that are not made of safety glass (e.g., not Pyrex) with aluminum foil.
  • Liquids
    • Fill containers only half full. This will allow for liquid expansion.
    • leave caps loose or cover with foil to allow steam penetration and prevent explosion.
  • Do not combine strong oxidizing material (such as dry hypochlorite) with organic materials (such as paper, cloth, or oil).
  • Do not autoclave flammable, combustible, reactive, corrosive, toxic, or radioactive materials.
  • Inspect glassware for cracks. Do not autoclave cracked or compromised glassware.
  • For bagged items, loosely tape or tie closed. Leave an opening for steam to penetrate the bag.
  • Place empty flasks or tubes horizontally to prevent air pockets.
  • Do not mix solid and liquid materials in one load.
  • Make sure items are not touching the walls or floors of the autoclave

Choosing the Correct Autoclave Cycle

Gravity (or “fast exhaust)

  • During this cycle, steam displaces the air in the chamber, and the force of gravity causes the heavier air to exit the chamber via the sterilizer drain. Once the chamber fills with steam, it holds it at a set pressure and temperature for a fixed time. At the end of the cycle, a valve opens, and the chamber rapidly returns to atmospheric pressure. Drying time may also be added to the cycle.
  • Suitable for Non-porous items such as glassware (for example, media bottles, beakers, and flasks),

Liquid (or slow exhaust)

  • This cycle prevents sterilized liquids from boiling over by releasing the steam slowly at the end of the cycle. This allows the pressure in the containers to reach equilibrium without causing the liquid to overflow.
  • Suitable for media, buffers, water,

Pre-Vac

  • Air is actively removed from the chamber by a series of vacuum and pressure pulses.  Suitable for surgical instruments in sterilization pouches, pipette tip boxes

Setting an Appropriate Time for Sterilizing and Drying

Always follow your lab’s standard operating procedures. 

These guidelines are recommended sterilization times

  • Dry goods- 30 minutes of sterilization plug 20 minutes of drying time.  Drying time might need to be extended for enclosed items
  • Liquids (bottles with vented caps)
    • Less than 500 mL- 30 minutes
    • 500 mL to 1L- 40 minutes
    • 2 L to 4L- 55 minutes
    • More than 4L- 60 minutes

There are other items to factor into your sterilization time including viscosity of the solution, container material, packing density (allow space between each bottle) and location in the autoclave.

Liquid cycles may take additional time than the set sterilization time due to slow charge and exhaust.

You should let liquids stand at least 10 minutes after the cycle is finished before opening the door.

Unloading an Autoclave

  • Make sure that the chamber pressure has returned to Zero before opening door
  • After donning PPE, open the door slowly and carefully (stand behind door)
  • Allow steam to be released and wait at least 30 seconds before opening door completely.
  • Wait for autoclaved materials to cool down before retrieving them (especially liquids)

Autoclave Issues

If the autoclave is not working properly, contact CBC facilities at cbc-facilities-management@osu.edu 

Please contact facilities if you encounter any of the following:

  • No steam
  • Cracked or compromised gasket
  • leaking water
  • insufficient jacket pressure
  • insufficient temperature
  • items are still wet even after a dry cycle