Basic Safety Rules SV vademecum Safety information Version 1.2 17.04.2013 GUIDELINES FOR THE MANGAGEMENT OF BIO- MEDICAL WASTE AT THE SCHOOL OF LIFE SCIENCES Table of contents 1. General Information... 2 1.1. Preliminary remarks... 2 1.2. Responsibility... 2 1.3. Basic rules... 2 1.3.1 Genetically modified organisms... 2 1.3.2 Infectious organisms from risk group 2 (BSL2 organisms)... 2 1.3.3 Genetically modified organisms... Erreur! Signet non défini. 1.3.4 Infectious organisms from risk group 2 (BSL2 organisms)... Erreur! Signet non défini. 1.4. Waste collection methods... 2 2. TREATMENT & ELIMINATION OF LIQUID WASTE... 2 2.1. Inactivation by Heat... 2 2.1.1. Cell culture medium... 3 2.1.2. Bacterial & yeast culture medium... 3 2.2. Treatment with chemicals... 3 2.2.1. Warning... 3 2.2.2. Inactivation of liquids containing biological materials... 3 2.2.3. Examples of products for treating biological liquids... 3 2.2.4. Products for the decontamination of surfaces... 4 Sometimes not efficient enough against non-enveloped microorganisms and rarely efficient against spores.... 4 Active against non-enveloped viruses. Good cleaning agent for AAV.... 4 3. Treatment & elimination of solid waste... 4 3.1. Inactivation by heat (autoclave)... 4 3.1.1. Autoclave bags... 4 3.1.2. Autoclave bins... 4 3.1.3. Pipets... 4 3.1.4. Sharps... 5 BIO_BSR_MBM_Waste_E_1 2.docx Page 1 of 5
1. General Information 1.1. Preliminary remarks This procedure is valid for BSL1 (biosafety level 1 or P1) and BSL2 (P2) biological waste. Elimination of BSL3 (P3) biological waste is described in the document BIO_BSF_MBM_Waste_P3.pdf. Animal carcasses or animal products are eliminated through a separate channel. Please refer to CAV rules and guidelines for more information and to the document BIO_BSR_MBP_Waste_Carcasses_[E]_1.0.pdf 1.2. Responsibility Each research unit, core facility or service is responsible for the proper sorting out of their biological waste. Only the bags or containers that have been prepared according to the procedure described below will be accepted for disposal. 1.3. Basic rules Always separate liquid waste from solid waste. 1.3.1 BSL1 non-infectious and non-genetically modified bio-medical waste Waste from bio-medical activities (cell cultures, non-infectious and non-genetically modified biological samples, enzymes, non-toxic reagents, consumables, etc.) is eliminated through a special channel. Solid waste that was not into contact with infectious or genetically modified material is collected in bags with the specific color code:: [White with red stripes]. Bags should be handled with care, always wearing hand protection. It is forbidden to transfer materials from one bag to the other. Do not overfill the bags. Exception: Solid waste contaminated with toxic or highly hazardous chemicals must be sorted out and eliminated according to the rules described in the document Guidelines for the management of chemical waste at the School of life sciences. Liquid waste that was not into direct contact with infectious or genetically modified material must be sorted out according to their chemical properties. Refer to the document: Guidelines for the management of chemical waste at the School of life sciences. 1.3.2 GMO waste Waste from genetically modified organisms (GMO) must be eliminated safely: Liquid waste must be autoclaved or treated with chemicals before disposal. Solid waste from risk group 1 GMO must be autoclaved or incinerated. In the latter case, the incineration process must be approved by the DSPS and the national authorities, the storage and transport must be conformed to UN regulations and the packaging must follow the standard P621. Solid and liquid waste from risks group 2 & 3 organisms must be treated like infectious waste. 1.3.3 BSL2 & BSL3 infectious waste Waste from risk group 2 & 3 organisms (and above) must always be inactivated on site before elimination. A special procedure for BSL3 (P3) waste is in place. 1.4. Waste collection and treatment methods Solid infectious or GM waste is collected into Steri-Safe autoclave bags [Color code GREY or WHITE]. Infectious and GM liquids are collected into glass or PP Nalgene bottles with caps or equivalent containers. 2. Treatment and elimination of liquid waste Inactivation of liquid waste can be achieved by autoclaving or by treating the solutions with an appropriate chemical decontaminant. The choice between chemical treatment and autoclaving will depend on various factors: Volume: The efficiency of heat treatment will decrease when larger volumes of liquids are autoclaved. Typically, in the autoclave models commonly used in research laboratories, the inactivation T is not BIO_BSR_MBM_Waste_E_1 2.docx 2/5
systemically reached or maintained long enough if the vessel contains more than 4 L of liquid waste. Thus, for volumes of liquid waste 3 L, chemical decontamination is recommended. The presence of chemicals: Solutions should never be autoclaved when they contain solvents, bleach or radioactive substances. The presence of antibiotics: Antibiotics are destroyed with heat, not with chemical treatment. The presence of organic matter: some chemical will have a reduced efficiency in the presence of high amount of organic matter. Typically blood, feces should be autoclaved and not be treated with bleach or other chemicals, but should be autoclaved. 2.1. Inactivation by Heat 2.1.1. Cell culture medium Cell culture medium will be aspirated in glass or 4 L PP bottles with safety caps. (eg. Vacusafe bottles, or Nalgen PP 5 L with Transfer cap PP 83B 2X or equivalent). Bottles without safety caps (e.g. with aluminum foil covers) will not be accepted. Recover as much as possible of cell culture medium or washing solutions by aspiration. Never add bleach (or any solvent or corrosive agent) to waste culture medium that will be inactivated by heat. On collection days, tightly close the cap; decontaminate the outside of the container with a proper decontaminant (Biocidal+, EtOH 70%, Meliseptol, Biosanitizer etc.) Once treated by heat (autoclave), the medium is discarded in the sink. 2.1.2. Bacterial & yeast culture medium Bacterial culture medium (or from other microorganisms) will be decanted in 4 L Nalgen PP bottles with Transfer cap PP 83B 2X or glass or PP containers with a screwing cap. When ¾ full, close the bottle, decontaminate it on the outside with the proper decontaminant and depose it for collection. Bottles without screwing caps will not be accepted. 2.1.3. Waste media containing antibiotics, hazardous chemicals or high nucleic acid content Antibiotics: Prefer autoclaving. If heat treatment is not possible, antibiotic-containing solution must be eliminated as toxic chemical products. Other hazardous chemicals: When the cell or bacterial culture medium contains hazardous chemicals (bleach, cytostatic agents, PARP inhibitors, highly active pharmaceutical compounds, solvents, etc.), they must be eliminated as toxic chemical waste. Plasmids: prefer NaOH treatment for the treatment of solutions in which DNA coding sequences are of concern. 2.2. Treatment with chemicals 2.2.1. Warning Wear safety glasses when handling chemical products. When handling small volumes of contaminated solutions ( 500 ml), the liquids can be treated with a chemical decontaminating solution. Spoiled glassware or plastic ware has to be soaked in a decontaminating solution (e.g. Deconex 11, Gigasept AF. Lysetol, etc.). Culture medium containing toxic or special chemicals (e.g. heavy metals; nanoparticles) must be discarded according to the processes designed for the elimination of chemical waste products. 2.2.2. Inactivation of liquids containing biological materials A wide range of chemicals is known to have toxic effects (See Appendix 1). Efficiency will depend on the type of microorganisms (bacteria; cells, parasites, yeast), the stability and the concentration of the product, the time of exposure, the ph etc. (See Appendix 2). 2.2.3. Examples of products for treating biological liquids Bleach Bleach (also known as Javel or sodium hypochlorite) is a very efficient decontamination product. It is active against most of the microorganisms, but is completely inefficient in the treatment of spores. Bleach is also relatively corrosive, which reduces its use as a general decontamination product. Bleach is highly efficient at a final concentration of 2%. Higher concentrations (up to 5%) are only necessary for the treatment of solutions containing high levels of amino acids (e.g. solutions containing albumin). Some publications claim that as little as 0.5% bleach is active with highly sensitive organisms. Most bleach stock solutions available on the market are 14% concentrated. Dilute 4 to 7 x times ( 28 to 15 ml of 14% bleach for 100 ml of solution to be treated) to get the final active concentration. Let incubate overnight before discarding the bleach-treated solution in the sink. BIO_BSR_MBM_Waste_E_1 2.docx 3/5
Never autoclave a solution containing bleach. Sodium hydroxide Sodium hydroxide (NaOH) 0.1 M final (or 4% final) is recommended for the treatment of solutions where free nucleic acids are of concern. It is an alternative to bleach if one would reduce the amount of chloride released into waste water. Virkon Virkon (60% Potassium peroxomonosulfate, 20% Sodium Dodecylbenzenesulphonate, 10% Sulfamic Acid) is a very effective virucide, bactericide and fungicide. Ideal concentration is 1% final. Vesphene: Vesphene is a decontamination product specially developed to inactivate Mycobacterium strains. As Vesphene is highly toxic for the environment, its use will be restricted to laboratory areas where Mycobacterium strains are in culture. Lysetol: Lysetol is a glutaraldehyde-based product used mainly for the decontamination of surgical instrument. It is bacteriocidal, virucidal, fungicidal and active against spores. It can also be used to treat cell culture waste. Spoiled glassware or plasticware has to be soaked in a decontamination solution (e.g. Deconex 11, Deconex labclean, Gigasept AF forte, lysetol or equivalent), before being collected for cleaning. All decontaminant products have a short shelf-life and are unstable when diluted. Some might be corrosive. Please, refer to the manufacturers instructions of use before choosing a product. 2.2.4. Products for the decontamination of surfaces For example, products such as 70% Ethanol, Meliseptol rapid, Biocidal ZF, Mikfobac forte, Perform Q-plus, Biosanitizer or others are common decontamination solutions used for the treatment of surfaces (bench, instrument, floor cleaning, incubators etc.). They are (i) often not efficient enough for the treatment of contaminated solutions (dilution factor is too high), and (ii) sometimes not efficient enough against non-enveloped microorganisms and rarely efficient against spores. 70% Ethanol 60% to 95% alcohol solutions are common decontaminating solutions. Preferably use 70% EtOH or a mixture of 70 % EtOH with 5% isopropanol (100% EtOH is completely ineffective as a decontaminant). EtOH solutions are flammable: do not spray alcohol solutions towards an open flame or a hot surface. Protect your eyes and do not breathe aerosolized solutions. Biosanitizer Activated hydrogen peroxyde solution (< 2% H202 with anionic surfactant compounds), nontoxic (does not contain volatile organic compounds), non-corrosive and nonflammable. Is active against non-enveloped viruses. Good cleaning agent for AAV. 3. Treatment & elimination of solid waste 3.1. Inactivation by heat (autoclave) 3.1.1. Autoclave bags Biological waste that needs to be autoclaved is collected in Steri-Safe-Bag that can be ordered through the SV faculty shop (Réf. Nbre 221 800 in SV magasin). [Color code GREY or WHITE]. The bags must show the Biohazard warning sign. When full, tightly close the bag, decontaminate the outside of the bag with a proper decontaminant (see point 2.2), tag it with the group bar code and dispose for removal. 3.1.2. Autoclave bins Bacterial plates, yeast cultures, C. elegans plates, semi-solid waste (feces, rodent s droppings) are collected in solid autoclave bins with a sealed melting cover. The bins are distributed by the SV-IN washing/waste core facility. 3.1.3. Pipets Plastic pipets: Place used pipets in specially design soft plastic carton containers. When cartons are full, trash them into the appropriate waste bag (BSL2, BSL3, bio-medical). BIO_BSR_MBM_Waste_E_1 2.docx 4/5
Do not put back pipets into their protection foil before trashing them The bins are distributed by the SV-IN washing/waste core facility. Glass pasteur pipets: Soak them into a decontamination solution (Deconnex, Virkon, lysetol, etc.). When treated, eliminate like glass material. 3.1.4. Sharps Sharps, needles, glass debris, microscope slices, cover slips that need to be autoclaved are collected into Yellow Sharp Boxes showing the Biohazard warning sign. Once ¾ full, the box is tightly closed, sealed, decontaminated on the outside and handed to the collection staff. Non-contaminated sharp materials (needles, slices, etc.) have to be collected in similar Yellow Sharp Boxes, without the Biohazard warning pictogram. Once full and tightly closed, they can be put into the bags dedicated to biomedical research waste. BIO_BSR_MBM_Waste_E_1 2.docx 5/5