College of Biological Science - Standard Operating Procedure
Autoclave
Date:
Revision:
10-Jan-2008
New
Prepared by:
Supersedes:
m Rush, G Humphreys,
A Doane
n/a
Purpose:
To provide instruction on proper use of autoclaves by research laboratory staff and students.
Application:
All users of autoclaves within the College of Biological Science should be familiar with the
procedures described below. This procedure has been written for the common use STERIS
Model SV-120 Scientific Eagle© Century Series Prevacuum Sterilizers. Autoclaves for shared
use are located in:
Integrative Biology
Molecular & Cellular Biology
SCIE 3402A
SCIE 1402
SCIE 2402
SCIE 3222B
SCIE 4402B
SCIE 4223B
Contacts: marie Therese Rush x58379
Contacts: Leanne Dickie x52660
Greg Humphries x58650
Karen Ingram x 52660
Safety Precautions:
o All operators must receive training on the safe operation of the autoclave prior to using
the equipment. Training may be delegated to a qualified individual, but it remains the
responsibility of the supervisor to ensure their personnel are adequately trained.
o Always wear suitable personal protective equipment. Closed toed shoes, lab coats and
insulated gloves must be worn when handling autoclaved materials. When a splash
hazard exists, face shields and aprons must be worn.
o Never attempt to sterilize flammable materials. The sterilizer is not designed for
flammables and this may cause a fire or explosion.
o When sterilizing liquids, use liquid cycle only and do not agitate containers of heated
liquids. Jarring containers can cause hot-bottle explosions. Place containers on carts or
trolleys lined with paper to avoid shattering of glass.
o Use only heat resistant glass (Type I borosilicate glass, Pyrex), or heat resistant plastics
(e.g., polycarbonate, PTFE, many polypropylene plastics) that are suitable for sterilization.
Ensure all lids have loose, vented closures to prevent pressurization/vacuum.
o Sterilizers, racks, and materials will be very hot after processing. Stand back from the
sterilizer when opening the door, and allow materials to cool before unloading.
o Immediately clean and spilled material or condensate from the floor to prevent slips or
falls.
o Keeps hands and arms out of the door opening when closing the sterilizer.
o Report any problems with the equipment, including unexpected noises, vibration, or
smells to the contact above.
o Place any sharps to be autoclaved in a designated sharps container.
o Do not attempt to open the door when a cycle is in process, or in the event of an alarm
(warble tone). An alarm may indicate the chamber has filled with scalding water.
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Certification:
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In compliance with Technical Standards and Safety Authority (TSSA) requirements and
University Policy 851.07.09, autoclaves are inspected annually. Certificates are posted in
the area of the equipment.
Preventative maintenance is performed regularly by an external contractor and recorded
in equipment log book.
Procedure:
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Preparation of Materials
o Confirm all materials and containers are safe to autoclave.
 Oils, waxes, some plastics, flammable materials, radioactive materials,
and samples containing solvents or substances which may emit toxic
fumes must not be autoclaved.
 All glassware and plastic to be autoclaved must be heat resistant.
Glassware should be inspected glassware for cracks. Examples of heat
resistant plastics include polycarbonate, PTFE, and many types of
polypropylene.
o If any biohazardous materials are to be autoclaved, ensure they are labeled
appropriately place in suitable container or autoclavable bag.
o For loose, dry materials, place in a container loosely covered with aluminum foil
or bag/wrap with an autoclavable, steam penetrable material.
o For liquids, ensure containers are less then 2/3 full and lids/covers are fully
loosened.
o Any sharps must be placed in a designated sharps container.
o Place items to be autoclaved in plastic or metal secondary containers for ease of
handling and to capture any spills. Secondary containers must have sufficient
volume to contain the contents.
o Apply thermal indicator tape to materials
Loading Autoclave
o Ensure personal protective equipment and clothing is adequate. You must wear
closed toed shoes and a lab coat and gloves when loading the autoclave. If the
sterilizer has been recently used, wear insulated gloves to protect against
accidental contact with hot surfaces.
o Step on foot pedal to open autoclave door.
o Place materials to be sterilized into the autoclave. Do not overload the sterilizer.
Steam must circulate in order to effectively sterilize the contents.
o Step on foot pedal to close autoclave door.
Operating Autoclave
o From touch panel, select the appropriate cycle time for sterilization. There are 6
cycles to choose from (see Table 1 below)
o For liquid sterilization, use Cycle 3 and refer to Table 2 for recommended
sterilization times.
o When cycle is complete, a message will display on the screen
o Never attempt to open the door while the autoclave is in operation.
o If you must abort, press the red box on the touch screen. Do not immediately open
the autoclave, as there may be residual steam after the cycle has aborted. After
waiting for at least a minute, stand well back and cautiously use the foot pedal
open the door slightly to release residual steam. Close door, then reopen fully.
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Table 1 – Autoclave Cycles
Cycle 1: HRD 20
- Prevac
- 15 psi (for 121°C)
- 20 minute sterilization time
Cycle 2: Sharps
- Prevac
- 26 psi (for 132°C)
- 60 minute sterilization time
- 5 minute dry time
Cycle 3: Liquid Adjustable (Lqd_adj)
- Adjustable cycle – operator
selects
- 20 minute sterilization time
(adjustable)
- Temperature and dry times
adjustable
Cycle 4: Liquid20
- Standard Liquid cycle
- 121°C
- 20 minute sterilization time
Cycle 5: Pip Tip
- Prevac
- 15 psi for 121°C
- 20 minute sterilization time
Cycle 6: Dry_adj
- Adjustable cycle – operator
selects
- 20 minute sterilization time
(adjustable)
- Temperature and dry times
adjustable
Table 2 – Recommended Liquid Sterilization Times for 121°C
Largest volume in any single container
Minimum recommended sterilization
time
25 minutes
75 ml
30 minutes
250 ml
40 minutes
500 ml
45 minutes
1000 ml
50 minutes
1500 ml
55 minutes
2000 ml
55 minutes + 10 minutes per litre
>2000 ml
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Unloading Autoclave
o Wear a lab coat, insulated gloves and closed toed shoes when unloading. A face
shield and splash apron should be worn if a splash hazard exists.
o Stand back from the door, and depress the foot pedal. Keep back from the door as
residual steam may be released..
o Allow sterilized material to stand for 10 minutes before unloading. For large
loads, items may need to stand for an additional 10 minutes. Do not remove caps
or agitate containers of heated liquids.
o Remove items from autoclave and place in an area which clearly indicates the
items are ‘hot’.
o Allow items to cool to room temperature. Place material on cart lined with paper
or paper towel. This will prevent shattering of glass which may occur if it is place
directly on metal.
o Clean the autoclave. Do not use chloride-containing solutions as these may
corrode the metal components of the autoclave.
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Autoclave Log
o Entries must be placed in the log books each time autoclaves are used. These
records are used for maintenance/service schedules and reporting of incidents,
accidents or faults. Log information must include date used, name of operator &
research lab, cycle used, and cycle parameters for cycles 3 & 6.
Sterility Assurance
o Quality control programs for sterility assurance are coordinated by the autoclave
contacts and are not the responsibility of typical operators.
Contingency Plans
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Equipment Malfunction
o If you encounter smoke or steam escaping the unit or water flooding the floor,
press the Emergency Stop (red button) and immediately leave the room. Contact
the appropriate person for the autoclave and post a notice on the door.
o If there autoclave does not operate exactly as expected, but the malfunction does
not appear to put people or property in danger, abort the cycle using abort button
on touch screen. Place notice on autoclave indicating that it is out of service.
Contact the appropriate contact person (see above) to arrange for investigation or
repair.
Incident Response
o If any injury occurs, seek first aid, and if necessary dial 2000 and request
emergency medical assistance.
o Should clothing become saturated with hot water or steam, remove it immediately
and cool the affected area in cool water.
o Place a notice on the autoclave indicating that it is not to be used until the
accident has been investigated.
o All incidents must be reported using the University of Guelph Injury/Incident
Report Form, available at:
http://www.uoguelph.ca/ehs/Forms/incidentreportform.pdf. The supervisor of the
individuals involved must be notified, and form must be submitted to the
department. The completed form must be faxed to EHS within 24 hours.
Spills Cleanup
o Spills may occur from boil over or breakage of containers. Do not attempt to
operate the autoclave until the spill has been cleaned up.
o If a spill occurs, prevent the spill from spreading and wait until both the autoclave
and the material inside have cooled to room temperature. Do not attempt to clean
a spill when it is hot. Clean affected area and dispose of waste appropriately. Any
broken glass must be placed in the designated pail.
o The operator of the equipment is responsible for spill clean up.
o Record the spill in the autoclave log book.
Applicable Policies & Regulations:
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University of Guelph Safety Policy 851.07.01
University of Guelph Safety Policy 851.07.09
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