1. Autoclave Training Guide York University Department of Occupational Health and Safety Suite C, East Office Building, 4700 Keele St. Telephone : 416-736-5491 www.yorku.ca/dohs/biosafety Revised: 16.November 2010

2. Autoclaving Training Outline Autoclave Overview Autoclave Hazards What can be Autoclaved How to Autoclave  Cycle selection  Proper containers  Proper loading/unloading Autoclave Performance Indicators

3. Steam Sterilization Pressurized device that uses heat, steam and pressure to achieve sterilization or decontamination

4. Autoclave Overview Typically operated at 121°C, 15psi, for 15-45 minutes. Allows the heating of liquids above boiling point. Uses moist heat (steam) to increase efficiency of sterilization. Heat is used to coagulate proteins, which destroys microorganisms and any potential biohazard.

5. Autoclave Hazards Tremendous pressure from steam in chamber provides explosive potential. High temperatures creates potential for burns and scalding. Potential exposure to hazardous fumes Inadequate decontamination allows for the potential of biological hazards to contaminate personnel and the environment.

6. Autoclave Hazards

7. What you CAN autoclave Biological waste that can be autoclaved:  Microbial stocks and cultures  Items contaminated with such waste: petri dishes, eppendorf tips, pipettes, gloves, paper towel Autoclaving is also used for:  Items for sterilizations such as; glassware media, aqueous solutions

8. What you CAN’T Autoclave DO NOT Autoclave:  BIOMEDICAL WASTE Including human anatomical or blood waste, animal anatomical or blood waste, cytotoxic waste, or any waste in contact with these waste products (including sharps waste).  RADIOACTIVE WASTE  HAZARDOUS CHEMICAL WASTE This includes anything contaminated with a toxic, volatile, corrosive, or mutagenic chemical (e.g.) bleach, formalin, glutaraldehyde, ethidium bromide Check MSDS beforehand

9. Autoclaves used to treat biohazardous waste at York Lumbers Room 120A Farquharson 227A Chemistry Building 447

10. How to Autoclave 1. Preparing your items for autoclaving 2. Loading the autoclave 3. Choosing the cycle settings 4. Unloading the autoclave 5. Aborting Autoclave

11. Autoclaving: Preparing your items Sign into log book  Keeps track of autoclave use for maintenance records Use personal protective equipment!  Eye protection  Heat resistant gloves  Lab coat

12. Autoclaving: Preparing your items Use a primary container  Container comes into direct contact with the contaminated or non-sterilized material or fluid  Do not fill more than 75% of holding capacity

13. Autoclaving: Preparing your items Use a primary container (cont’d)  Must NOT be a tightly sealed container (might explode)  Primary container must permit heat (steam) penetration Loosen screw caps or use self venting caps Cap open containers with aluminum foil or muslin If using plastic waste bags, leave a small opening Loosen screw caps Small opening

14. Autoclaving: Preparing your items Use a secondary container  Used to contain any spills  The sides of the secondary container must be sufficiently high to contain any spill that may occur  Tray MUST be autoclave safe For liquids add water

15. Autoclaving: Preparing your items Use temperature sensitive tape  Will indicate that high temperature has been achieved  Will not prove that decontamination or sterilization was successful  Will assist in keeping track of autoclave and non autoclaved items BeforeAfter

16. Autoclaving: Loading the autoclave NEVER autoclave liquid loads using a solid load cycle  liquids will boil over Be cautious if autoclave was recently used  Rack may be hot  Use heat protective gloves Make sure tray is locked on dolly  Rack may slide out unexpectedly if not locked

17. Autoclaving: Loading the autoclave Farquharson:  Tighten door so that prongs are fully extended  If noise and/or steam escapes, abort the cycle and tighten the door more Lumbers  Keep the door button pushed until the ready prompt is on the screen

18. Autoclaving: Which cycle to use Autoclaves run 3 types of cycle programs The type of cycle depends on what is being autoclaved: Liquid/Slow exhaust* For autoclaving liquids * Prevents liquids from boiling over Solid/Gravity* Best for unwrapped solid items (ie glassware) Solid/Vacuum* Best for wrapped solid items Note: for Farquharson, use Solid/Gravity for wrapped, solid items

19. Autoclaving: Choosing a Cycle For Liquids:  20 mins / litre of liquid, 5 mins per additional litre For Solids:  Glassware (empty): 15 mins  Instruments (utensils): 30 mins  Biohazardous Garbage: at least 30 mins per full bag Use biological test strips to optimize duration

20. Autoclaving: Proper loading Autoclave biohazardous waste separately Do not over load primary or secondary containers  Allow for sufficient steam penetration  Do not fill more than 75% to allow expansion without overflow Do not pack or compress contents Unautoclaved Autoclaved (regular) Autoclaved (due to being compressed & sealed)

21. Autoclaving: Unloading autoclave Use PPE Wait for autoclave to state END CYCLE before opening door When opening, stand away from door opening  Make sure no one is standing by door opening Farquharson: turn door knob slowly and open door slightly to allow steam to escape Lumbers: make sure your hands are not above the top vents

22. How to Autoclave – Unloading autoclave Make sure rack is locked on to dolly Unmark any biohazard signs that may be seen on waste bags Dispose of waste in proper bins Please put back heat resistant gloves for other users Keep autoclave doors shut when not in use

23. How to Autoclave – Aborting Only qualified personnel should attempt to troubleshoot an autoclave Farquharson  Large autoclave: push abort button  Small autoclave: need to manually advance through the autoclave cycle Lumbers  Push abort button

24. Autoclave: Performance Indicators How to know if autoclave is functioning correctly: Physical - Annual testing by certified technician - Pressure, Temperature, Cycle times, recorded on paper Chemical - Heat sensitive autoclave tape - Not an indicator of successful sterilization, useful to keep track of autoclaved and unautoclaved items Biological - Tests ability of autoclave to sterilize effectively - Bacillus stearothermophilus spore strips often used because they are resistant to steam sterilization. - EZ Test (SGM Biotech) (Fisher Sci #29801 074) - 3M Attest Rapid Readout Biological Indicators - Steris Verify Integrator Laminated and EO Integrators

25. Autoclave Issues/Concerns? Contact: Your Supervisor Farquharson and Lumbers: Jane Grant, Biology, Lumbers 130, x22657 Debbie Freele, Biology, Lumbers 115, x22655 Prof. Roger Lew, Biology, Farq 230, x66114 Chemistry Building: Philip Johnston, Chemistry Building, x33119 York Biosafety Officer: Debbie Kolozsvari, DOHS, x44745

26. Autoclave Training Completion of in-class autoclave training is included with the Biosafety Training session. For practical, hands-on training, contact your supervisor.