OHS026
Safe Work Procedure
Faculty/Division
Medicine
School/ Divisional Unit
SOMS
Document number
Initial Issue date
Current version
SOMS_IIR_SWP16
10.3.10
1.1
Current Version
Issue date
10.3.10
Next review date
10.3.11
The Writing Safe Work Procedures Guideline (OHS027) should be consulted to assist in the completion of this
form.
Safe Work Procedure Title and basic description
Title: Safe use of CO2 incubator (Sanyo and Forma Scientific) incubator in room 524A CIIR
Description:
Description: How to operate CO2 incubator for culturing cell lines in culture plates and flasks using liquid
media. The CO2 incubator maintains temperature and humidity as well as appropriate CO2 level in the
incubator necessary for optimal cell growth.
This SWP is to ensure the correct operation of incubators to help reduce the likelihood of contamination in
the incubator and to the cell lines and the necessary decontamination procedure.
Associated risk assessment title and location: Risk Assessment for operating the CO2 incubator (16_RA
CO2 Incubators) and risk assessments for the individual experimental protocols using cell lines. Also view
the manufacturers’ instructions for hazards/MSDS for all media and its components.
Describe the activity or process
Personal protective equipment (PPE) consisting of full sleeve laboratory grown (blue) andlatex
gloves must be worn before starting work
Note
1. The CO2 incubators have been installed by the supplier as per manufactures instructions. The
incubators are connected to an external CO2 manifold via a reticulation system. DO NOT
TAMPER WITH any plastic hosing connecting the incubator to the CO2 reticulation system.
2. The temperature of the incubators is generally set at 37oC and at CO2 concentration of 5% and
are ready for use.
3. One Sanyo incubator (lower incubator on right hand side against the middle bench) has been set
at reduced CO2 conditions. DO NOT USE this incubator unless your cells require these conditions
for growth.
General operation of an incubator that is ready for use
1. Check the temperature and the CO2% reading on the incubator prior to opening the outer door
2. Open glass inner door and place flasks gently on the shelf making sure that liquid media is not
tipped towards the lid. With plates take care not to tip sideways so that no liquid spillsfrom the
plate.
3. Close the glass door and the outer door gently.
To set up operation of an incubator that has been switched off
Note
The incubator needs to be switched on day before it is required so that the temperature and CO2
concentration stabilise to the set conditions.
1. Check that the incubator is connected to CO2 reticulation tap and that this tap is in the closed
position
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Current Version: 1.2, 15/08/2007
2.
3.
4.
5.
6.
7.
8.
9.
Check that CO2 incubator is connected to the power supply and switch on.
Turn on the incubators using the on/off button located on outside of the incubators.
Set the required temperature using the up/down arrow keys after selecting the temperature
Open doors and check water tray in the bottom of the incubator has sufficient sterile water for
correct humidity maintenance.
Close doors and leave undisturbed for 6-8 hours for temperature to stabilise to the required set
temperature.
Open the CO2 tap gently to start the gas flow
Set the CO2 concentration in the incubator by using up/down arrow and let equilibrate overnight
for stable CO2 atmosphere.
Next day follow general operating procedure when placing cells inside.
List all resources required including plant,
chemicals, personal protective clothing and
equipment, etc
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


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Latex Gloves should be worn when handling the flasks and plates
Laboratory Long sleeve gown.
Carbon Dioxide, 70%(w/v) Ethanol, 4% bleach
Risk assessments for the individual experimental protocols using cell lines.
Tissue culture flasks and plates
CO2 incubators List potential
List potential hazards and risk controls including
specific precautions required
Hazard: Possible CO2 leakage. - lower CO2% reading on the incubator display panned and activated
audible alarm
Risk control: Check that Incubator doors are closed securely.
Risk Control: Check that CO2 connection is sealed safely.
Risk Control: Check that CO2 supply is adequate, by checking the CO2 manifold outside Room 542A.
If CO2 cylinders connected to the manifold need to be replaced with full cylinders see lab manager or
person in charge of tissue culture room
Risk Control: If all the above have been checked and are in order, a lower CO2 reading can also occur if
the temperature sensor or temperature display are faulty or if the instrument is incorrectly calibrated.
Arrange for a Service / calibration for the instrument.
Hazard: Biological spill.
Risk control: Make sure the plates or flasks have no breaks before use and place plates and flasks of
cultured cells carefully in the Incubator.
Risk control: Contact laboratory manager immediately for advice on how to clean up the spill with use of
4% bleach and fill out an accident report.
Risk control: gown and gloves
Hazard: Electrical malfunction
Risk control: Electrical protection switch, electrical tagging, annual service and maintenance.
List emergency
List emergency shutdown instructions
In the event of an electrical hazard, turn off immediately at the power.
If there is CO2 leak try to turn of cylinders at the manifold if safe to do so and evacuate area.
Contact laboratory manager or OHS representative ist
List clean up and waste disposal requirements
If there is a biological spill in the incubator, clean and disinfect as follows:
1. Use 70% (w/v) Ethanol for cleaning. Do not use abrasives or caustic solutions of bleach as they
can cause pitting and rusting of the interior of the incubator and affect the CO2 sensor
2. Carefully remove flasks/plates from the incubator and check for contamination. Wipe the outside of all
Flasks /Plates with 70% (w/v) Ethanol and place in a clean cabinet.
3. Remove incubator-shelves, shelf supports and humidity tray and clean individually with 70% (w/v)
Ethanol
4. Clean incubator chamber and doors, inside and out with 70% (w/v) Ethanol.*
5. Replace shelf supports and shelves. Make sure all shelves and supports are completely dry before
returning to the incubator.
6. Replace the humidity tray with sterile water
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Current Version: 1.2, 15/08/2007
7. Allow the temperature and CO2 levels to stabilize before returning flasks to the incubator.
*Note After washing as per points 1-4 it is preferable to autoclave all shelves and removable
brackets to destroy fungal spores
List legislation, standards and codes of practice used
in the development of the SWP
General: ANZCWMIG
EMIAA
Chemical: NSW OHS Act and Regulation
AS/NZS 2243.2
Biological: AS/NZZ 2243.3
AQIS
GTA&R
Supervisory approval, training, and review
Supervisor:
Signature:
Plant custodian: Eva Fiala-Beer
Signature
List competency required – qualifications, certificates, licencing, training - eg course or instruction:

Laboratory induction

Course OHS - OHS awareness for employees

Course PC2 - PC2 bio training

Couse HAZ - Working with hazardous substances
SWP review date: April 2011
Responsibility for SWP review:
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Safe Work Procedure
Uncontrolled document when printed
Date Effective: 01/01/2007
Current Version: 1.2, 15/08/2007