LBL02001

Laboratory Induction

Author: S. Thompson	Approved by:	SOP No. LBL02001
Signed:	Signed:	Effective from:
Date:	Date:	Last edited:

1. Purpose

This describes the laboratory induction procedure for all new users of the laboratory facility. It is a requirement that this be completed by all new laboratory users and their confirmation of it's requirements recorded by signing the appropriate training sheet.

2. Scope

This applies to and is required by all users of the laboratory. If there are any questions please contact the biological safety officer (BSO).

3. Responsibilities

Users of the laboratory by being inducted by the biological safety officer are hereby acquiring the responsibilities of individuals working in a laboratory environment. They should be made aware of and agree to respect the hazards and risks that are or could potentially be present within a biological laboratory working to containment level one standards. Further, for all procedures carried out by the laboratory user they should be aware of all other SOPs that address the kind of procedures they wish to carry out. Also, the principal investigator for each project is required to carry out a risk assessment of the specific procedures that may be involved in their proposed project. Particular attention should also be paid to the containment, handling and disposal of genetically modified organisms (GMOs) and agreed with the genetic modification safety committee (GMSC). In case of any doubt please ask the BSO for further information.

The current BSO is Samantha Thompson. Contact: bioscisam@gmail.com , tel 07720677678.

4. Related documents

All SOPs relavent to further work to be carried out.

5. Definitions

BSO – Biological Safety Officer
GM – Genetically Modified
GMO – Genetically Modified Organism
GMSC – Genetic Modification Safety Committee

6. Procedures

6.1 Laboratory Access Control The lab can be accessed using the number lock or using the card sensor to unlock the door, if your access card (such as an oyster card) has been registered. Access control to the laboratory area is to ensure that only trained and responsible individuals can make use of the laboratory facilities of which this risk assessment is primarily concerned. Those wishing to access the laboratory should only do so after going through and understanding the safety and operational factors highlighted in this document. This induction process should be conducted by the Biological Safety Officer (BSO) also any further relavent LBL Standard Operating Procedures (SOPs) should be read and understood where this pertains to the work the user desires to carry out. Lab users should be concious of the safety requirements and skill level necessary when working with reagents and equipment needed in their particular projects and should consult the SOP documentation and/or BSO if unsure. Once completed the user and their confirmation will be added to the list of authorised lab users by the BSO. The status and content of the SOP framework is the responsibility of the Biological Safety and Genetic Modification Safety (where relevant) Committees and they should be be informed of the nature of the work any users of the laboratory wish to undertake. The immediate point of contact is the BSO. For those starting new projects in the laboratory full details of the requirements of principal investigators, in particular risk assessments for new projects can be found in [LBL SOP # ..]

6.2 PPE, Biosafety and Contamination Risks The kind of biological organisms with which we typically work in the laboratory include i) non-pathogenic strains of E. coli (such as DH5 alpha), ii) cellulose producing gram-negative bacteria such as Gluconacetobacter, iii) non-toxic Dinoflagelate species, iv) brewer’s yeast (Saccharomices cereviscae) and, v) algal microorganisms from non-toxic environmental samples.

These organisms do not themselves require working containment level 1 (CL1) standards however it is still considered good laboratory practice to minimise exposure of any laboratory user to direct contact with them and prevent any biomaterial from being unintentionally carried out of the laboratory. Further, those organisms which do fall into the classification of CL1 requirements such as those strains genetically modified by transformation for example E. coli by heat shock method and of Gluconacetobacter by electroporation, should have extra care taken to ensure that any surplus material is destroyed in the kill bin and by autoclaving of waste. Personal Protective Equipment (PPE) suitable for working with these kinds of organisms is provided and consists of lab coats, goggles and gloves which the BSO will indicate to the new inductee. If suitable PPE is not available to hand the BSO should be informed immediately before commencing any practical work. To ensure that any contamination of genetically modified biomaterial and microorganisms does not occur, laboratory users should be aware of the correct handling and disposal procedures required. They should be made aware of typical requirements during laboratory safety induction and this knowledge should be reinforced during individual project risk assessments. Generally, all microorganisms should be handled while using gloves, eye protection and protective clothing to prevent material coming into contact with the user’s skin or eyes. This is both to protect the user and to eliminate the risk of transporting biomaterial out of the laboratory after work is completed. Gloves can be disposed of in the bins provided. Containment level 1 biomaterial should be deactivated before disposal by autoclaving and immersion in bleach in the ‘kill bin’ underneath the sink. The deactivated waste in the kill bin can then be autoclaved and then diluted further and disposed of down the drain. These handling and disposal procedures are explained later in the SOPs but are highlighted here to communicate the important message that biosafety and protection of users of the laboratory is of the highest importance.

6.3 Microwave The microwave is a familiar device to probably all laboratory users and the standard precautions apply when using it. In particular, the handling of hot liquids and glassware may present the risk of burns and as such thermally resistant gloves are provided. Of note is also the fact that one of the primary uses of the microwave is heating of agarose for the casting of electrophoresis gels. User’s should be aware that previous, possibly less careful, operators may have used the microwave in conjunction with biocontaminants and/or hazardous chemicals (as described later) and as such operation of the microwave using gloves and goggles is recommended in any case. [associated SOPs]

6.4 Spectrophotometer The spectrophotometer is a device that measure the absorbance of a given liquid sample at various wavelengths of light. A full description of it’s usage is given in the SOPs [SOP]. User’s should be aware that this system can generate light of significant intensity in both ultraviolet and infrared wavelengths. While most of the optical path and the bulbs required to generate this light is contained within the system, the user may come into contact with this in the sample measurement compartment of the device. As such it should be operated only with the sample measurement comparment fully closed.

6.5 PCR Machine The Polymerase Chain Reaction (PCR) machine, also known as a thermalcycler, is used to amplified specific sections of template DNA either for diagnostic or molecular cloning purposes. The heating block and underside lid of this machine will typically reach temperatures in the region of 95 to 100degC in a typical amplification program and caution should be exercised when placing and removing samples. Information regarding the current state of the machine including current program and temperature is displayed on the LCD on the front panel of this machine. Full information regarding the use and programming of the Techne thermal cycler and it's use in various technical can be found in the SOPs and technical documentation. [SOPs, docs]

6.6 Electrophoresis Power Supply Unit The Power Supply Unit (PSU) used to supply a current to the electrophoresis tank (described below) is capable of producing dangerously high voltages and currents. It is typically set to the levels required for DNA electrophoresis in 100mL 1 - 2% w/v agarose gels. Caution should be exercised when plugging in and handling electrodes due to risk of electric shock. Use of a residual current device is recommended when using this or any other electrical device in the laboratory that may pose a risk to operators from electric shock. This instrument is also often used in conjunction with Ethidium Bromide based procedures (see below) and should be operated using gloves.

6.7 Electrophoresis tank and Ethidium Bromide area The electrophoresis tank is located within the electrophoresis/Ethidium Bromide area of the laboratory bench. It is advised any work carried out within this area is manipulated separately from any work done in other areas. Any consumables used when handling Ethidium Bromide such as gloves and tips should be disposed of before handling anything outside of this area to prevent contamination of the wider lab area with the potentially toxic chemicals used in DNA staining. Separate pipets designated for ethidium bromide work and labelled as such should only be used for molecular biology work in this area and these pipets should not be used anywhere else. A full description of how to set up and run agarose gels for DNA electrophoresis is described in the SOPs [SOP].

6.8 UV illuminator Gel electrophoresis separates DNA by size within an electric field. Separation of varying sizes of DNA fragment within an agarose gel allows visualisation of the various sizes of DNA fragment contained within a sample when nucleic acid binding stains are used. The typical stain used is Ethidium Bromide which is a fluorophore that is excited within the UV spectrum (with excitation maxima under 300nm) and emission within the visible spectrum. This presents two main risks to the user: i) As a DNA intercalating agent Ethidium Bromide is potentially carcinogenic and should never be allowed to come in contact with the user’s skin, caution should therefore be applied throughout the entire DNA electrophoresis procedure from gel preparation, through electrophoresis and then subsequent gel visualisation. This is therefore also carried out in the electrophoresis/ethidium bromide area of the laboratory bench. ii) UV light can be damaging to exposed surfaces of the body and to the eyes. In extreme circumstances or prolonged use this can lead to carcinomas or eyesight damage. It is therefore advisable that eye protection be used by all people present within the laboratory when UV gel visualisation is taking place and that any possible contact from UV radiation on the user’s body while manipulating and viewing their gel be minimised through adequate PPE and controlled use of the UV light source. [SOP for gel vis]

6.9 HEPA flow cabinet/area The HEPA filtered laminar flow unit allows us to work in sterile air in order to prevent contamination of our work such as petri dishes and broths with other microorganisms. Proper usage of the laminar flow area is described in the SOPs. It should be noted that the laminar flow functions in such a way as to protect the user’s work rather than the user and so caution still be taken by the user to maintain the aseptic conditions of anything used within the flow area and in disposal of consumables to assist in containment of biomaterial. See SOPs on aseptic technique. [SOP].

6.10 Hot plate/Steam cooker/steriliser/autoclave The sterilisation unit [autoclave?] is used to destroy any potential microorganisms that might contaminate media, reagents and consumables to be used aseptically in micro and molecular biological procedures. High temperature and pressure is used to kill contaminants and the main risk to the user is from the heat of the metal pressurised unit during sterilisation and any vented steam. The system itself is unpowered and is heated using the hot plate which itself becomes hot enough to cause serious burns. Thermal gloves are available for handling the autoclave and material processed by it. It is recommended that the autoclave and contents are allowed to cool for a while before handling. The correct procedures for using the autoclave/steriliser to prepare media and destroy GM waste are described in the SOPs. [SOPs]

6.11 Sink area The sink area should remain clear. All glassware should be kept clean and out of the way.

6.12 Chemicals and Storage As per chemical list, and MSDS by chemical [SOPs appendix A]

6.13 Incubator While the incubator itself does not pose a high risk due to typically operating in the temperature required for the culture of mesophiles. care should be taken when moving samples to and from the incubator as per handling biocontaminants mentioned previously. Many procedures described later in the SOP documentation indicate the need for incubation and details can be located there. [SOPs]

6.14 Centrifuges The laboratory has a number of centrifuges available for use. The Jouan is a larger device which can take 50ml tubes and larger containers if fitted with buckets and can spin up to 10,000 rpm (although documentation and instrument panel indicate higher rpm may be possible this particular unit appears to have a limit of 10,000. The Henle and MSE microcentaur can spin up to higher rpm with smaller samples. These centrifuges will not operate in their normal state without a closed lid preventing the user from coming into contact with the moving parts. There are also two blood centrifuges which spin in the region of 1000 to 2000 rpm while much slower can be operated with open lids which is strictly advised against for obvious safety reasons but primarily exposure of dangerous moving parts to the operator and a risk of spray from poorly sealed liquid samples. All centrifuges should be cleaned after use for the purpose of biosafety, containment and equipment maintenance. SOPs described later which require centrifugation provide further details on usage and settings. [SOPs]

6.15 Refrigerator, Freezer and Sample Storage

The refrigerator maintains samples and reagents in the area of 4degC such as bacterial plates, and reagents for transformation. Due to the presence of biomaterial in the refrigerator caution should be exercised to prevent contamination of the user. The freezer contains longer term bacterial stocks and enzymatic reagents and similar caution should be taken. Specific details for the handling of the various chemicals and reagents stored in the freezer and refrigerator can be found in the chemicals and storage list [SOP appendix A]. Specific SOPs also cover the storage and recovery of frozen bacterial stocks and procedures requiring the usage of frozen and refrigerated reagents. [SOPs]

7. Resources

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