STANDARD OPERATING PROCEDURE
Procedure:
SDS-PAGE
School/Department:
SOP prepared by:
School of Biological Sciences
Nick Coleman (SMB) edit M Joseph
Section 1 - Personal Protective Equipment
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Lab coat or lab gown
Nitrile or latex gloves
Safety glasses at stages where squirt/splash/spray risk is present
Proper enclosed footwear
Hair tied back if long
Dust mask when handling SDS powder
Section 2 – Potential Hazards + Safety precautions
1. Before you begin, consider if pre-cast gels are commercially available for your
procedure. These gels reduce many of the hazards mentioned below.
2. Acrylamide used to make gel. Potential for inhalation, skin/eye contact or
ingestion with harmful effects. Acrylamide is a potent neurotoxin.! Wear gloves!
3. SDS is harmful by inhalation, ingestion, skin or eye contact. SDS is an irritant. Wear
gloves & dust mask when handling powder.
4. TEMED used for making gel. Potential for inhalation, ingestion, skin or eye contact
with harmful effects. TEMED produces a strong stench and is a corrosive alkali.
TEMED is also flammable. Wear PPE as above.
5. APS used for making gel. Potential for inhalation, ingestion, skin or eye contact
with harmful effects. APS is a strong oxidizer and is corrosive. Wear PPE as above.
6. Mercaptoethanol in the buffer used for denaturing protein samples. Potential for
inhalation, ingestion, skin or eye contact with harmful effects. Mercaptoethanol
produces a strong stench, and is toxic. Wear PPE as above, and handle
concentrated mercaptoethanol in the fume hood.
7. Electrophoresis. Potential for serious electrical shock or electrocution due to
leaking chamber, faulty or corroded electrode cables, or faulty power supply.
Ensure all equipment is in good working order, and that lid is always on
electrophoresis tank when in operation.
8. Protein samples. May be recombinant in nature and/or may have other
hazardous properties (eg. toxins). Follow approproiate guidelines and work in a
PC2 lab if required.
9. Chipped glass plates. Potential for causing lacerations or deep cuts to skin. May
also result in leaking of unpolymerised acrylamide solution (refer to above for
hazard description). Discard any chipped or damaged plates.
10. Loading samples via needle syringe. Potential for needle stick injury. Handle with
care, do not re-sheathe needles.
11. Dispensing Methanol (present in staining and destaining solutions). Potential for
exposure via inhalation and direct contact with eyes and skin. May be readily
absorbed through skin with possible harmful effects. Methanol primarily affects
the central nervous system with symptoms of headache, nausea, and dizziness.
Potential fire hazard. Wear PPE, dispose of as hazardous waste.
12. Dispensing Acetic Acid: (present in staining and destaining solutions) Potential for
exposure to highly corrosive vapours, and for direct contact of corrosive solution
with eyes and skin. May result in irritation, redness and pain, burning and
Issue date: 20/11/12
Review date: 21/11/13
Page 1 of 4
ulceration. Wear PPE.
Section 3 – Procedure
1. Read and understand this SOP and the risk assessment for SDS-PAGE, along with
any MSDS sheets
2. Put on PPE as described above
3. Know the location of spill kits, eyewashes, safety showers before starting work
4. ONLY purchase and use pre-made acrylamide solutions - this removes the
significant hazard of inhaling the toxic powder form.
5. Avoid ALL contact with acrylamide solutions or polyacrylamide gels.
- do not handle gels without gloves on !
6. LIMIT acrylamide work to designated areas where possible. Coat work bench
with absorbent bench paper prior to working with acrylamide to prevent
contamination of non-disposable surfaces. Change this paper layer frequently
(eg. weekly or monthly, depending on work intensity)
7. USE barrier/filter tips or disposable plastic transfer pipettes when aliquoting
acrylamide solutions to help prevent contamination of pipettes.
8. Solutions containing acrylamide, and the resultant polyacrylamide gels must be
disposed of as hazardous chemical wastes to room 225, not down the sink
9. Dispense TEMED, mercaptoethanol and glacial acetic acid in fume hood to
minimize stench. Keep containers tightly closed. Keep ignition sources well clear
of TEMED vapours – 3 metres is advisable. Do not inhale TEMED,
mercaptoethanol or acetic acid vapours, or allow contact with eyes or skin.
10. Inspect electrodes and cables for defects prior to use. DON’T use any
electrophoresis equipment that you suspect has an electrical fault or is
damaged – in this case, report item to supervisor/ lab manager/ safety officer for
electrical testing, repair or replacement. Ensure electrophoresis chamber has a
secure LID which prevents accidental contact with the electrified buffer solution
11. If the protein samples have come from recombinant organisms or clinical
specimens, these need to be handled in a PC2 lab, using PC2 methods, and any
wastes disposed of according to PC2 guidelines (see related risk assessments on
risk group 2 microbes and clinical samples). Protein samples may have other
associated risks eg. toxins – always handle with gloves, and dispose of excess via
autoclaving or similar (not down the sink). Know the details and potential risks of
the proteins you are working on before you start handling them.
12. Use containers with secure lids when staining and destaining gels, and decant
solutions slowly and carefully to prevent splashing.
13. Ensure area around electrophoresis chamber is free of spills or leaks PRIOR and
DURING the running of your gel
14. For detailed procedure for making and running SDS-PAGE gels, consult
instruction manual of electrophoresis apparatus, or practical manual from
appropriate course.
Section 4 – Disposal / Spills / Incidents
1. Depending on the nature of the spill, clean up as described in SOPs for
Biohazard Spills, Flammables, Corrosives, or Toxic Substances. Any acrylamide
spills must be cleaned up immediately and thoroughly using first dry paper towel,
then wet paper towel, and the towels disposed as hazardous waste.
2. Depending on the nature of the materials used, dispose of wastes as described
Issue date: 20/11/12
Review date: 21/11/13
Page 2 of 4
in SOPs for Biohazard Spills, or Chemical Wastes
3. Any large spills of hazardous materials (>1L) or incidents resulting in injury must be
reported to your supervisor immediately and via the online incident report form
within 24 h. Near misses (dangerous situations not leading to an incident) should
also be reported
Section 5 – Repairs / Certification / Validation
1. Ensure electrophoresis equipment is kept in good repair. Consult the
manufacturer or the SMB service centre if in doubt
Section 6 – Relevant Material safety data sheets
1. Consult MSDS for acrylamide, TEMED, APS, SDS, acetic acid
Section 7 - References
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Essential: Risk assessment for SDS-PAGE
As needed: Risk assessment and SOP for Flammables, Toxic chemicals,
Disposing of chemical wastes
SOP Training Confirmation
By signing below, these individuals confirm that they have read and
understood the SOP, and agree to always follow the instructions in this
SOP when performing this procedure.
Position
Supervisor
employee / student
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Name
Signature
Date
WHS Committee Approval
Representative: A. Prof Frank Seebacher Chair Safety Committee
Signature: ........................................................ Date: .....................................
Issue date: 20/11/12
Review date: 21/11/13
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