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Safe Work Method Statement Acrylamide Gel Electrophoresis SWMS NUMBER RISK ASSESSMENT NUMBER RISK ASSESSMENT Medium DATE CREATED / REVIEWED 15.07.13 Employees Involved in SOP Creation Dr. Tim Chataway, Angela Binns CONTACT PERSON Dr. Tim Chataway Personal Protective Equipment Required Laboratory coat. Covered shoes Mask to be used when weighing the chemical SDS. Gloves – to be worn throughout entire procedure. Eye protection as required Before work Commences Users must be trained and training documented. Refer to the SDS for each chemical Only use commercially made Acrylamide (DO NOT make your own use only pre-made) Read and understand the procedure. Hazards Hazardous substances Electric shock – electrophoresis power supply. Lacerations Needle-stick injury Eye damage 1. Reagents Acrylamide solution Tris buffers SDS solution Ammonium Persulphate TEMED Sample buffer The hazards are listed on the SDS for each chemical and are listed on Chemwatch. Essentially, gloves must be worn throughout this procedure. If the gloves are contaminated with one of the chemicals, the gloves must be changed or rinsed thoroughly in water before proceeding. A mask must be worn when weighing the chemical SDS. Solutions Acrylamide Acrylamide solutions should not be made from dry powder. Acylamide is a cumulative neurotoxin and the dry powder is hazardous. There are commercially available pre-made solutions which should be used. Tris buffers Tris buffers do not pose any significant hazard. However, they need to have their pH adjusted using strong acids or bases. Gloves and eye protection must be worn. Read the SOP for using a pH meter. Equipment Electrophoresis involves the use of power supplies which produce lethal levels of electricity. However, the risk is negligible if safe procedures are followed. Ensure that the leads are inserted correctly, all fluids are kept away from the power supply and the electrophoresis tank is assembled with its lid in place before attaching the power supply. Procedure Clean glassware with Extran solution Remove adherent acrylamide with razor blade Assemble plates and spacers into gel pouring apparatus Make up acrylamide solution of required percentage Degas solution Add catalyst, pour solution into plate apparatus, allow to set disassemble plates, insert plates into electrophoresis tank, add upper and lower tank buffers Prepare samples in SDS loading buffer, heat at 95 deg C in heating block Load samples into gels Apply voltage (200Volts) Remove gels from tank, separate glass plates, place gel in tray for staining, add Coomassie Blue Destain with methanol/acetic acid solution Risk This procedure involves potential exposure to toxic chemicals Eye injury or laceration if plates are dropped Finger laceration Eye injury or laceration if plates are dropped Exposure to toxic chemicals Precautions Gloves must be worn for all steps in this procedure Eye protection, wear gloves, covered shoes Use single sided blade only, apply at an oblique angle Eye protection, wear gloves, covered shoes gloves Explosive release of glass/acrylamide Use glassware designed for vacuum use Burnt fingers, lid pops off if heat too high Do not touch block, ensure correct temperature, label heating block as being hot to prevent others from accidental burns Load samples with 10 uL pipette Ensure leads connected correctly, correct buffer and fluid levels, lid in place on tank before applying voltage Eye protection, gloves Needlestick injury if using needle and syringe Electric shock Injury from glass Eye injury from acetic acid Use with care, eye protection