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NU Prepared by Number Date HSE HSE section Approved by The Rector HMSRV-26/01 Page 1 out of 6 01.12.2006 Replaces 15.12.2003 Hazardous activity identification process Unit: IBT Molgen Date: 27.08.2013 Updated 02.02.2016 SHA Participants in the identification process (including their function): You (M.student/PhD/PostDoc/researcher), Supervisor (Professor) Student/employee/guest Name (Capital letters) Supervisor/leader Signature Name (Capital letters) Signature Short description of the main activity/main process: Molecular genetics and microbiology ID nr Activity/process Responsible person Laws, regulations etc. Existing documentation All lab work 1 Acrylamide gel, making of Procedure must be written before acrylamide is used 2 Agarose gel, making, running, Geldoc Manual 3 Alginate isolation, purification and nonradioactive alginate assay Protocols 4 Antibiotic-stock solution, make and use of Lab-rules, relevant MSDSs 5 Work with liquid N2 Regulation for filling and transport. NTNU HSE guidelines for work with chemicals and gasses Existing safety measures Comment Nitrile gloves are to be used for all labwork, vinyl gloves can be used in order to protect the sample (but does not offer the same protection to the user). Nitrile gloves, protective eyewear, ventilation hood Nitrile gloves, protective eyewear (with UV filter), face shield when needed. Gelred used for staining or alternatively Gelgreen. Blue and white screens. Nitrile gloves, protective eyewear when using NaOH, Fume hood when using ether Procedure not currently performed Blue- and white screens stops UVirradiation when mounted properly Risk evaluation according to which chemicals are used Nitrile gloves, handle powder only inside fume hood Glasses, thick glowes. Easily removed footwear. Avoid shoes with wide opening to avoid spilling N2 into the shoes. Suitable containers for transport. Training necessary. See following point regarding transport of N2 NU Prepared by Number Date HSE HSE section Approved by The Rector HMSRV-26/01 Page 2 out of 6 01.12.2006 Replaces 15.12.2003 Hazardous activity identification process 6 Filling and transport of liquid nitrogen 7 Autoclave 8 Bacteria class 1 and recombinant bacteria Manual Operating instruction Approval: (GMM regulation)(”Godkjenning av laboratorier og anlegg for innesluttet bruk av genmodifiserte mikroorganismer” (08.12.10 Helsedirektoratet) 9 Bacteria class 2 Approval letters from arbeidstilsynet (11.06.10 and 27.04.07). 10 Recombinant bacteria class 2 Approval ”Godkjenning av laboratorier og anlegg for innesluttet bruk av genmodifiserte mikroorganismer” (08.12.10 Helsedirektoratet) Glowes, face protection, tank and carrier for transport, easily removed suitable shoes (not rubber boots). O2-meter by the filler. Do not take the elevator with the N2 (l)-tank. Thermo-resistant gloves Eye protection lab, Instructions posted regarding not opening autoclave before temperature <80 °C, not overfilling bottles or closing bottle lids completely (to avoid broken bottles). Autoclave accessible. Inactivation of gene manipulated bacteria in contaminated material and waste. Lab coat mandatory. Lab bench surfaces resistant to water, acid, alkali, solvents, disinfective agents, decontaminating agents and easy to clean. Transport between labs only in closed containers. Good microbiological practice as defined in http://www.lovdata.no/for/sf/ho/to20011221-1600-018.html According to risk assessment: Risk assessments: “Novel anti-infective agents from marine bacteria (19.04.07)” and “Sintef project no: 80117000” (14.011.08) and “Risikovurderinger og rutiner, arbeid med mikroorganismer i smitterisikogruppe 2” (28.05.09). Autoclave in building. Inactivation of gene manipulated bacteria in contaminated material and waste. Lab coat and gloves mandatory. Lab bench surfaces resistant to water, acid, alkali, solvents, disinfective agents, decontaminating agents and easy to clean. Transport between labs only in closed containers. Restricted access to lab. Good microbiological practice Room D4-106 (SINTEF). Separate rules. Contact SINTEF for further arrangement Room D4-106 (SINTEF). Separate rules. Contact SINTEF for further arrangement NU Prepared by Number Date HSE HSE section Approved by The Rector HMSRV-26/01 Page 3 out of 6 01.12.2006 Replaces 15.12.2003 Hazardous activity identification process (http://www.lovdata.no/for/sf/ho/to20011221-1600-018.html). Biological hazard sign on door. Measures to minimize aerosols. Control of vectors (closed windows, no food I lab, general awareness of possible vectors), 11 12 Bioanalyzer Use of open flames – (e.g. sterilization with Bunsen burners) 13 Cellcracker (Mini bead beater) General lab work 14 Refer to the appropriate section in lab-rules Bunsen burner must not be left burning Manual Laboratory handbook NTNU, MSDSs, EcoOnline system, signature for training/introduction to lab rules Refer to respective kit protocols No risk Disposable inoculation loops will be tested as an alternative to glass. No risk Safety rules according to risk assessment Use gloves and eye protection during steps including NaOH. Use fume hoods for procedures containing chloroform, phenol or if it is indicated in the kit manual Use nitrile gloves or eye protection if the kit contains irritating fluids or proteases 15 DNA/RNA isolation and purification 16 DNA/RNA kit for enzymatic modifications or clean up from reactions Manuals 17 Electroporator, use of Manual 18 Environmental samples for cultivation and DNA isolation, preparation of Risk assessment (projects) 19 Enzyme-assays (i.e. betalactamase-assay, phosphoglucomutase-assay, epimerase assay...) Relevant protocols According to MSDSs of relevant chemicals 20 Ethanol/dry ice bath Lab-rules 21 FPLC, running of Refer to instrument instructions. Precaution at instrument. Use thermo-resistant gloves and eye protection, loose-fitting (e.g. rubber) boots should not be used. According to MSDSs of relevant chemicals Avoiding contact with high voltage by following user instructions from manual. According to risk assessment No risk when used as stated in manual. Risk evaluation by specific project Potential damage to eyes and frostbite NU Prepared by Number Date HSE HSE section Approved by The Rector HMSRV-26/01 Page 4 out of 6 01.12.2006 Replaces 15.12.2003 Hazardous activity identification process 22 Luciferase measurements, Promega Luciferase Assay kit Manuals According to MSDSs of relevant chemicals No risk 23 Luminometer and fluorometer, use of Manual According to MSDSs of relevant chemicals No risk 24 Mammalian cells, class 2 Risk evaluation and procedures must be written for each experiment/new organism. 25 Preparation of medias for growing bacteria Recipe According to MSDSs of relevant chemicals 26 Mutagenesis using a mutagenic agent (e.g. Etylmethanesulfonate, methylethanesulfonate, nitrosoguanidine, UV) Experimental procedure Risk assessment and approval from project leader Nitrile gloves, fume hoods, requires written procedure and risk assessment for the experimental procedure (typespecific) including detoxifying waste and how to warn and protect others working in the laboratory. Description and risk assessments must be written and approved by the responsible project leader (signature). Approval is mandatory not only for each experimental procedure, but also for each new person to perform that procedure 27 Nano-drop measurements of DNA/RNA Manual 28 Northern and Southern blotting Manual 29 PCR PCR-manual Room D4-106 (SINTEF). Separate rules for HTS/cell lab. Contact SINTEF for further arrangement. Hepatitis B vaccine offered. No risk No risk According to MSDSs of relevant chemicals. Gloves and lab coat, work in a fume hood if MOPS and/or formaldehyde is used. Usual precautions when working with UV light and DNA staining. Fume hood when DMSO is added No risk NU Prepared by Number Date HSE HSE section Approved by The Rector HMSRV-26/01 Page 5 out of 6 01.12.2006 Replaces 15.12.2003 Hazardous activity identification process 30 pH-adjustments Manual 31 Protein isolation and purification 32 Proteingel-gels. Running of CBS – manual http://cbsscientific.com/pdf/DCXInstructionsNov201 0.pdf Sambrook and Russell, Molecular Cloning: A laboratory manual. section A.8.42 33 Pulse field electrophoresis Manual Operating instruction 34 Radioactivity work including use of scintillation counter http://www.nt.ntnu.no/ibt/innsidadokumentlager /HMS/Veileder%20for%20arbeid%20med %20radioaktivt%20materiale%20IBT.pdf Risk evaluation and procedures must be written for each experimental procedure Operating instruction scintillation counter. 35 Centrifugation (Sorvall + table) Manual Operating instruction 36 Sonication Manual Operating instruction Nitrile gloves, eyewear, shoes/shoe bags with protection against acid/base spill, sufficient ventilation (hood/”cap”) According to MSDSs of relevant chemicals Nitrile gloves (e.g. protection against acrylamide), eyewear, staining and de-staining in fume hoods Protection against voltage (lid etc.) to avoid electrical shock. Avoid contact with de-staining reagents (methanol/acidic acid). According to relevant MSDSs. Protection against voltage (lid, gloves etc.) to avoid electrical shock. Gloves, eye protection, lab coat, hood. Testing of work area for contamination after work. Accurate balancing, accurate attachment of rotor, not exceed maximal G-forces for each type of tube Ear protection. Pregnant women are not allowed to stay in the room during sonication. Sonication cabinet with sound absorbing material. Shut doors of the room where sonication is taking place, and post warning sign on door. No risk (confer bacteria, sonication and chemicals used when applicable) Department rules state that the local radiation- protection coordinator must preapprove any users of radioactive compounds and experimental procedures must also be approved. The scintillation liquid used may contain dangerous solvents NU Prepared by Number Date HSE HSE section Approved by The Rector HMSRV-26/01 Page 6 out of 6 01.12.2006 Replaces 15.12.2003 Hazardous activity identification process 37 Supercompetent or electrocompetent cells, making of Eye protection, gloves and protective shoes/shoe bags when handling liquid nitrogen or ethanol/dry ice bath. Shoes must be easy to take off in case of spill into (never use rubber boots) 38 Transformation 39 Ultrasound waterbath, use of 40 UV/Vis Spectrophotometer 41 Ventilation hoods, use of Operating instruction 42 Washing machine, use of Manual 43 Waterbaths and shaking waterbaths, (heat and water quality) Manual 44 Western blotting No risk Operating instruction Ear protection. Pregnant women are not allowed to stay in the room during sonication, close the door and post warning sign. Avoid skin/eye exposure to UV light. Never disassemble the instrument. Follow instructions/manual when changing light bulb. Opening minimized when not in use, keep the hood-opening small when in use (for efficient protection). No risk No risk Timer on water baths. Large water baths (> 1 L) are not to be used to boil small samples. Put up notices if heating the water baths at temperatures > 70°C. Chemicals. Avoid contact with buffers (e.g. methanol). NTNU Prepared by Number HSE/KS HSE section Approved by The Rector HMSRV2603E 04.02.2011 Page Replaces 01.12.2006 7 out of 6 Risk assessment Unit: IBT -molgen Line manager: Kjetil Rasmussen Date Date: 27.08.2013 Updated 02.02.106 SHA Participants in the identification process (including their function): You (M.student/PhD/PostDoc/researcher), Supervisor (Professor) ID nr Activity from the identification process form 2 Agarose gel electrophoresis Potential undesirable incident/strain Likelihood Consequence: Risk : value Likelihood Human Environment Economy/ Reputa (1-5) (A-E) (A-E) material tion (A-E) (A-E) Unknown A Unknown Long term health risk from GelRed 1 A B (unknown effect) UV-light: skin or eye damage 2 2 B A A B 2B 5 Work with liquid N2 Frostbite 2 C A A B C2 5 Work with liquid N2 Suffocation 1 E A A E E1 6 6 Filling/transport of liq N2 Filling/transport of liq N2 Frostbite due to spurt of liquid N2 Low oxygen atmosphere 2 1 B B A A A A B D 2B 1D Comments/ status Suggested measures Gelred and Gelgreen is said not to penetrate cell membranes, and thus should not act as mutagen even if it is DNA-binding. Gloves also minimize the risk for exposure. UV damages on unprotected skins/eyes if instructions not followed Follow regulations for filling/transport. Oxygen measurement? Follow regulations for filling/transport. Oxygen measurement? O2-meter by NTNU Prepared by Number HSE/KS HSE section Approved by The Rector HMSRV2603E 04.02.2011 Page Replaces 01.12.2006 8 out of 6 Risk assessment tank/filler. Do not ride the elevator together with the N2 (l)-tank! Will not happen if instructions are followed Rapid pressure fall due to opening the 2 autoclave to soon my cause hot liquid burns on eye or skin Bacteria class 1 and recombinant Release of GMO to environment 2 bacteria Bacterial infections C A A B 2C A A A B 2A 9 Bacteria class 2 Bacterial infections. Release of infective, pathogen bacteria to environment. 2 C B A C 2C 10 Recombinant bacteria class 2 Bacterial infections. Release of infective, pathogen gene manipulated bacteria to environment. 2 C B A C 2C 12 Bunsen burners Skin burns 3 B A A A 3B 12 Bunsen burners Initiation of fire. 3 C A B A 3C 14 General lab work Deviation from established safety rules and -routines -- -- -- -- -- 15 DNA/RNA isolation and purification Exposure for dangerous for health chemicals: Phenol-chloroform mix C A A B 7 8 Autoclave Date 2 2C Low risk because of strict regulations and good lab practise. Low risk because of strict regulations and good lab practice. New rules how to handle burners+ ethanol baths already installed. New rules how to handle burners + ethanol baths already installed. No injuries requiring more than simple first aid in these laboratories for the past 5 years, the present routines seem sufficient Phenol-chloroform mix requires the work in a ventilation hood only. All waste should be placed in NTNU Prepared by Number HSE/KS HSE section Approved by The Rector HMSRV2603E 04.02.2011 Page Replaces 01.12.2006 9 out of 6 Risk assessment unknown 18 Environmental samples for cultivation and DNA isolation, preparation of Bacterial infections, production of 1 toxic/infectious compounds from cloned DNA. Usage of hazardous compounds (e.g. chloroform) for DNA isolation. 20 Dry ice/ethanol Eye injury. Frostbite 1 B 22 FPLC, running of Leakage of buffer 23 Luminometer and fluorometer, use of 24 Mammalian cells, class 2 26 28 Date a special box for hazardous materials. In general mild 1unknown injuries, that can be avoided by present routines and sensible carefulness during lab-work. 1B Eye-protection and Gloves for handling the Dry ice A A C A A B 2 See A MSDS A Depends on injury 1A Possible exposure health injurious chemicals. 2 A A A A 2A See MSDS for relevant chemicals Exposure to biological agents, infections. 2 D A A D 2D Mutagenesis using a mutagenic Exposure to mutagens agent (e.g. Etylmethanesulfonate, methylethanesulfonate, nitrosoguanidine, UV) 1 E A A E 1E Applying aseptic procedures. Good microbiological practices. Probably too small amounts to ever damage the environment. Because of the known risk of cancer, these experiments are always conducted with extreme caution, and using gloves and fume hoods. Northern/Southern 1 C A A A 1C Exposure to injurious chemicals. Northern blot may involve formaldehyde and NTNU Prepared by Number HSE/KS HSE section Approved by The Rector HMSRV2603E 04.02.2011 Page Replaces 01.12.2006 10 out of 6 Risk assessment 29 PCR 30 pH-adjustments 32 Protein gel-eletrophoresis 33 Pulse field electrophoresis acrylamide which are toxic, thus fume hood and protective clothing must be used. Due to the use of toxic chemicals special disposal methods are required. Exposure to DMSO when used 1 Date A A A A 1A Exposure to corrosive chemicals: Skin and 2 eye damage Electrical shock 1 B A A B 2B E A A C 1E Electrical shock and depending on method used for DNA preparation :) 1 E A A A 1E 33 Exposure to hazardous chemical: phenylmethanesulphonyl fluoride (toxic, corrosive) 1 C A A B 1C 34 Radioactivity work including use Health injury as result of radiation of scintillation counter (external, internal as result of uptake). Radioactive contamination of working environment. Centrifugation (Sorvall + table) Damage caused by loose rotor Sonication Damage to hearing (including foetus) Creation of injurious aerosols (infectious microorganisms, toxic agents) Supercompetent or Exposure of skin and eyes to extremely electrocompetent cells, making of cold liquid (nitrogen or ethanol) 1 E A A E 1E 1 1 C C A A B A C B 1C 1C 2 C A A B 2C 35 36 36 37 PCR-machine should be in ventilation hood when DMSO is added, use lab coat and gloves Eye-protection and lab coat Equipment for electrophoresis will only work when lid is closed, thus minimizing the possibility of suffering an electric shock. Use of gloves/fume hoods for handling of hazardous chemicals will minimize risk. Conducted with extreme caution, and using eyeprotection and NTNU Prepared by Number HSE/KS HSE section Approved by The Rector HMSRV2603E 04.02.2011 Page Replaces 01.12.2006 11 out of 6 Risk assessment Date gloves. 39 Ultrasound waterbath, use of Damage to hearing (including foetus) 1 C A A B 1C 41 Ventilation hoods, use of 1 C A A B 1C 43 Waterbaths and shaking waterbaths, (heat and water quality) Exposure to hazardous chemicals due to insufficient airflow (effects on local hood or other hoods) Electrical hazards, Burns 1 B B A A 1B 44 Western blotting Exposure to hazardous chemicals 1 See A MSDS A Depends on injury 1A Likelihood, e.g.: 1. Minimal 2. Low 3. Medium 4. High 5. Very high Consequence, e.g.: A. Safe B. Relatively safe C. Dangerous D. Critical E. Very critical Risk value (each one to be estimated separately): Human = Likelihood x Human Consequence Environmental = Likelihood x Environmental consequence Financial/material = Likelihood x Consequence for Economy/materiel Can be prevented by maintaining sufficient airflow Regular service by trained personnel. Heat-proof gloves must be used during handling of hot items. Hot surfaces must be labelled with warning signs. NTNU Prepared by Number HSE/KS HSE section Approved by The Rector HMSRV2603E 04.02.2011 Page Replaces 01.12.2006 12 out of 6 Risk assessment Date Potential undesirable incident/strain Identify possible incidents and conditions that may lead to situations that pose a hazard to people, the environment and any materiel/equipment involved. Criteria for the assessment of likelihood and consequence in relation to fieldwork Each activity is assessed according to a worst-case scenario. Likelihood and consequence are to be assessed separately for each potential undesirable incident. Before starting on the quantification, the participants should agree what they understand by the assessment criteria: Likelihood Minimal 1 Once every 50 years or less Low 2 Once every 10 years or less Medium 3 Once a year or less High 4 Once a month or less Very high 5 Once a week Consequence Grading Human E Very critical D Critical May produce fatality/ies C Dangerous B Relatively safe Serious personal injury A Safe Permanent injury, may produce serious serious health damage/sickness Injury that requires medical treatment Injury that requires first aid Environment Financial/material Very prolonged, non-reversible damage Prolonged damage. Long recovery time. Shutdown of work >1 year. Minor damage. Long recovery time Minor damage. Short recovery time Shutdown of work < 1 month Insignificant damage. Short recovery time Shutdown of work < 1day Shutdown of work 0.5-1 year. Shutdown of work < 1week The unit makes its own decision as to whether opting to fill in or not consequences for economy/materiel, for example if the unit is going to use particularly valuable equipment. It is up to the individual unit to choose the assessment criteria for this column. Risk = Likelihood x Consequence Please calculate the risk value for “Human”, “Environment” and, if chosen, “Economy/materiel”, separately. About the column ”Comments/status, suggested preventative and corrective measures”: Measures can impact on both likelihood and consequences. Prioritise measures that can prevent the incident from occurring; in other words, likelihoodreducing measures are to be prioritised above greater emergency preparedness, i.e. consequence-reducing measures. NTNU utarbeidet av Nummer Dato HMS/KS HMS-avd. godkjent av Rektor HMSRV2604 side 13 av 1 08.03.2010 Erstatter 09.02.2010 Risk assessment matrix CONSEQUENCE Risk assessment matrix NTNU Very serious E1 E2 E3 E4 E5 Serious D1 D2 D3 D4 D5 Moderate C1 C2 C3 C4 C5 Small B1 B2 B3 B4 B5 Very small A1 A2 A3 A4 A5 Very small Small Moderate High Very high PROBABILITY Color-coding in the risk assessment matrix Color Description Red Yellow Green Unacceptable risk. Measures must be implemented to reduce risk. Evaluation area. Evaluate whether measures need to be implemented. Acceptable risk. Measures may be considered based on other factors.