MHH Guidelines BSL 1 Genetic Laboratories

Medizinische Hochschule Hannover

Biosafety OE 0014

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MHH Guidelines BSL 1 Genetic Laboratories

Operational Procedures Biosafety Level 1 (BSL 1) Genetic Laboratory

pursuant to §12, Section 2, Gentechnik-Sicherheitsverordnung

I. Area of Application

MHH, Institute / Department …...

Rooms:

Project Leader(s):

Biosafety Officer: Dr. J. Mertsching

The Project Leader(s) and the Biosafety Officer are to be informed immediately of any breakdowns or deficiencies in the laboratory.

II. First Aid

First aid cupboards are situated in rooms …….. Dr. …… is responsible for first aid. In her absence, one of the project leaders or coworkers should be called to assistance.

Emergency Nos

Injuries and contaminations with microorganisms are to be reported to the first aider and the project leader without delay. The following immediate first aid measures should be taken:

-Upon skin contact, the contaminated area should be thoroughly cleansed with Sterilium.

- In case of injured skin respective are should be disinfected with Oktenisept.

-Following eye contact, the eye must be washed for at least 5 minutes with running water (Augen-Dusche).

-In the case of light burns and scalds, the affected area of skin should be immediately held under a continuous stream of water (20°C).

If, despite these emergency measures, symptoms persist help should be sought immediately from the Casualty Department. Every injury must be entered in the Injuries Book (room …..).

III. Instructions for Conduct of Work

- Recording of genetic engineering experiments

Before work commences the individual project leader responsible arranges for an experimental outline to be drawn up based on the MHH standard form (MS Word) and confirms its accuracy by signature. A hard copy of the outline is forwarded to the Biosafety Department where the data are transmitted by laser to an optical image recorder (WORM Medium). The application forms are an integral part of this outline. A copy of the outline remains with the project leader on site until experiments are concluded. Should the project leader leave, it must be clarified which experiments are complete and which will be pursued by another project leader.

- Safety Cabinets

The safety cabinets must be allowed to run for at least 20 minutes before use. The gas burners may only be ignited when the blower is switched on.

The work surface of the bench must be decontaminated after every use with ……(e.g. 70% ethanol or Bacillol AF). The following precautions must be observed when using alcohol-based disinfectants:

.the surface to be cleaned must not measure more than 2 m2

.the quantity of disinfectant must not exceed 50 ml per m2

.cleaning may not be carried out in the vicinity of a source of ignition

(e.g. naked flame of Bunsen burner)

.the surface temperature must not exceed 37°C

The blower must be switched off before beginning to clean the collection reservoir under the work bench. Otherwise, there is a danger that cleaning cloths will be sucked in thus destroying function.

- Autoclave

The autoclave may only be used after instruction from someone technically competent (responsible: Dr. ……). Biological waste must be sterilized for at least 20 minutes at 121°C and can be subsequently disposed of with the general laboratory waste. Biological waste is collected in autoclave bags in metal buckets and temporarily stored in the -4°C coldstore. There are specially designated bins for solid waste (petri dishes, etc.) and for liquid waste (cell culture remnants). Waste bins must not be filled more than three-quarters full.

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- Storage of Microorganisms

Permanent cultures may only be stored in marked containers in the coldstore (room ……), the –70°C freezer, the nitrogen tank or in the specially allocated storage facilities in cellar rooms ……. The cultures and the storage containers with associated storage list must be marked so as to be clearly identifiable with the organism, the vectors it contains and the lab group.

IV. Description of Organisms

Escherichia coli, Phage 

Various eukaryontic cell lines

Saccharomyces cerevisae

Mice

V.Dangers to Man and the Environment

According to the current state of scientific knowledge the aforementioned (micro-)organisms present no danger to human health or the environment. The E. coli stocks are typical laboratory bacteria derived from the security stock E. coli K12. Phage  infects exclusively E. coli stocks and displays no human or animal pathogenic characteristics.

VI. Protective Measures and Rules of Conduct

- A lab coat is to be worn in the laboratory.

- Lab coats must be removed before entering the kitchen. Special hooks for lab coats are located at the entrance.

- Mouth pipetting is prohibited. Mechanical pipetting devices are to be employed.

- Disposable gloves may only be worn at the lab bench. Gloves must not come into contact with doorhandles, telephone receivers or keyboards.

- Syringes and canulae should only be used when absolutely necessary. Canulae, scalpels and similar equipment must be deposited in the clearly marked containers.

- In all laboratory work care must be taken that no avoidable aerosols are generated.

- Glass objects for disposal and broken glass should be deposited in the specially designated containers.

- Laboratories should be kept clean and tidy. Only really necessary equipment and material should be left standing on the benches. Storage of supplies must be confined to the rooms, cupboards or shelving especially set aside for this purpose.

- Upon completion of work organisms must be correctly stored or disposed of in an appropriate fashion.

- Foodstuffs and tobacco articles may not be stored in the lab, storage must be confined to the kitchen ( room ……).

- Eating, drinking and smoking are prohibited in the laboratories.

1. Conduct in Emergency Situations

- Leakages

An uncontrolled release of microorganisms can occur during centrifugation of bacterial cultures when the centrifuge safety cups are filled too full. For this reason, the centrifuge safety cups for fixed-angle rotors may only be filled up to a level which does not permit overflowing during centrifugation. If a discharge of liquid still occurs, then the part of the centrifuge housing the rotor and the rotor itself must be disinfected with ……(e.g. Bacillol AF or 4% Dismozon (reaction time: ……).

When cultivating microorganisms in the oscillating water bath there is a danger that the screws holding the insets of the culture dishes in place may loosen causing the dishes to knock together and contaminating the water bath on the inside. The water must be autoclaved and the water bath cleaned using the above-mentioned disinfectants.

- Spills

If bacteria-containing liquids are spilled on lab benches or on the floor, the surfaces must be disinfected straightaway with Bacillol AF. Chemical spill kits are located outside the doors to rooms ……

- Fire

The fire alarm is located opposite to ……. Powder extinguishers are attached to the walls in the various laboratories.

- Escape Routes

The passageways to the staircases constitute the main escape routes. Should, in an emergency, these routes prove impassable, then windows must be used to escape.

VIII. Disposal of Liquid and Solid Waste

- As a rule, all equipment used for microbiological or genetic engineering experiments is to be treated as contaminated.

- Infected disposable objects must be collected in the specially reserved containers and finally autoclaved.

- Bacteria-containing pellets discharged during centrifugation (e.g. plasmid preparations) should be formed into a liquid suspension, transferred to a glass receptacle and finally autoclaved.

- Liquid waste must be collected in glass receptacles, temporarily stored in steel buckets and autoclaved.

- The pipette stands must be filled with the disinfectant ……(e.g. 0.6% Mucocit-P) to such a level that the glass pipettes are entirely covered.

- The autoclave must run for 20 minutes at 121°C in order to kill the bacteria. The persons named on the autoclave timetable are responsible for sterilization of contaminated waste.

- Plastic equipment which is not disposable and which cannot be sterilized with the autoclave must be disinfected in a …….solution (e.g. 1% Kohrsolex) for 1 hour.

- After sterilization in the autoclave, disposable material and liquid waste can be removed with the normal laboratory waste.

Hannover,……..2007………………………

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