Team:Elan Vital Korea/Safety

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WETLAB
-Safety-

Safety First!

Saving the world starts with saving ourselves. You cannot save the world unless you’re in good shape. We put safety on the top priority in any lab works. We have been informed accurately of the lab safety rules as well as bio-safety regulations at theinitial stage of the project in order to understand hazards and safety procedures. Also we received safety training from our instructor before commencing our lab work. Even though our project is included Bio-Safety Level 1, we know very well that there are still many risks that may be escalated into serious disaster. Furthermore, even when handling those risks that may not pose serious threats to the experienced scientists, we as young learners should be extra careful.

So, Let's begin by watching a video:

The rule of thumb is:

1. Start with reading Safety Questions for overview.
2. Follow Elan Vital Safety Rules all the time.
3. Before entering the laboratory, read GMS General Safety Rules and GMS Extra Rules again and follow them
4. If you are unsure of the rules or confused, ask lab assistant and/or instructor before doing anything.

About Biosafety

Team Elan Vital understands that biosafety is about preserving the health of people and the environment of the experiment alike ­ and we conducted our experiment with those factors in mind. Undergoing strict security guidelines, our team performed all experiments by adhering to all areas related to biosafety. From preventing mistakes to making sure that no matter what happened, safety issues would be reduced to a mild and manageable extent. We conducted all experiments under the appropriate protocols for the materials we would be dealing with, and we as a team made sure that all safety issues were taken care of and were constantly considered whenever we participated in an experiment.

Basic Information about our Experimental Safety Procedures

All experiments were conducted at Gachon University’s Molecular Biology Laboratory.

We followed the CDC code for Biosafety Level 1, designed for “work involving well-characterized agents not known to consistently cause disease in immunocompetent adult humans, and present minimal potential hazard to laboratory personnel and the environment” (CDC, Biosafety in Microbiological and Biomedical Laboratories 5th Edition). Safety precautions include:

• mandatory gloves
• goggles to protect eye
• no consumption of food or liquid, or any activity that could bring potential infection from the hand to the face, such as applying cosmetics
• minimize creation of splashes or aerosols
• Procedures for sharp materials, of which we only had to deal with broken glassware since we used no sharp materials
• For broken glassware: do not pick up or clean up directly with one’s hands, but remove using tools.
• separate waste receptacles for biowaste
• autoclave decontamination of all materials for bacterial cultures
• decontamination by disinfectants on hands and laboratory surfaces
• Protective laboratory wear (e.g. a lab gown)
• proper measures depending on the infectivity of our bacteria (in the case of E.coli, which is the only bacterial species we are using, very little)
• Protective eyewear if we had any hazardous materials, of which we had none Our Laboratory Facilities were also within requirements, with conditions such as:
  • Doors for access control
  • Sink to wash hands
  • Alcohol to disinfect hands
  • Easy cleaning environment
  • Bench tops that are resistant
  • Non­porous surfaces on chairs
  • Windows have screens
  
  
  

  Similar guidelines can be found here: http://www.cdc.gov/biosafety/publications/bmbl5/BMBL5_sect_IV.pdf

  Possible Threats and Risks

  The possible risks of our experiment are also very low, even if infection occurred in some sort of way. But just in case, we also minimized such risks by changing the bacteria we were working with, as well as in deciding what materials we were working.

  Our original project, which we modified in order to reduce chances of possible infection and harm to general public health, was on MRSA, a methicillin-resistant strain of Staphylococcus Aureus. Upon discovering that MRSA bacteria is a high dangerous bacteria that could harm the human body easily and is not permitted for experimentation by iGEM, we decided to remap our project, undergoing clear measures to make our project much more safe and secure. We did this by changing our subject bacteria from MRSA to E.Coli, which poses much less risk.

  We also considered safety when we used a safe strain of E.coli, called TOP10 (Invitrogen). It is a strain of naturally competent cell, without risk of infection or danger to humans and the environment. As mentioned, we never used any sort of dangerous biological agent, and the only bacteria we used was E.coli, since we tested antibiotic resistance in naturally competent cells.

We have conducted our experiments at Gachon University. Thus, we have always observed safety rules of the lab. We had Safety Education & Training Session before starting our lab work. Also, when we conducted our experiments, Gachon University staff supervised our work.

Gachon University General Lab Safety Rule

1. Labs should be maintained in clean, organized and orderly manner.
2. All participants including students and instructors must understand and observe safety rules and guidelines.
3. Before the lab activities, lab safety education must be taken in which contents of lab experiments, operation methods of lab equipment, rules and guidelines, and procedures in case of accidents must be properly understood.
4. Everyone must follow instructions of the responsible person in the lab and should not perform any activities which may cause any danger.
5. Everyone must understand emergency behavior guidelines. Emergency includes fire, emergency patient and other dangerous situation. Everyone must know the location of emergent phone calls, fire alarm, and power disconnecting switch, etc.
6. Everyone must understand how to use fire extinguisher and check the status of it.
7. Smoking, eating, drinking, chewing the gum are prohibited in the lab. Disposal of food and drink is also forbidden.
8. All participants must be properly dressed in the lab with the safety Consideration.
9. During the operation of the equipment and during the experiment, at least one person should attend and observe the situation.
10. Person who leaves the lab last must make sure that power is disconnected, flammable material is isolated, dangerous material is safely kept, water is disconnected, lab is cleaned and locked, and order of the lab must be double- checked.
11. Power wiring appliance must be properly installed to accommodate the experiments. Cables, and lines must be checked regularly to confirm that they are capable of handling the electrical load.
12. All equipment must be connected with the proper power Cable and must not be connected in a multiple power socket.
13. Flammable material must not be used near firearms, and must be kept far away from the electric socket, air conditioner and/or heaters.
14. Heaters and other heating devices that are not related to experiments must not be used in the lab.
15. Places where dangerous materials are kept must be identified with labels including the name of the material and the level of danger. (Explosiveness, spontaneous combustion, flammability, toxicity, water reactivity, oxidation, contagiousness, radioactivity, corrosiveness and other dangerous material.)
16. Chemicals must be kept in isolation in consideration of flammability, corrosiveness, and other chemical features.
17. When conducting chemical experimentation, tasting and smelling must not be allowed. Pipette using mouth must not be allowed.
18. Inflammable chemicals must be purchased in small amount when necessary.
19. Inflammable chemicals including acetone, oil, or gas must be kept in the place where ventilation is easy and access is infrequent.
20. Importing and exporting of high-pressure gas container must be done using transportation equipment.
21. When storing high-pressure gas containers, they must be safely and firmly fixed, isolated from flammable substances and inflammatory materials.
22. Experiments using flammable, explosive, toxic, or volatile gases or vapors must be carried out in a hood.
23. Reagents container or cabinets for storing reagents must be kept in cool, well-ventilated places without direct sunlight, and far away from fire and heat sources.
24. Experimental drugs shall be stored in a way not to be shaken by outside shock, and so that its storage bottles do not fall
25. Toxic materials shall be handled using protective films, or other safety tents to prevent damages from splatter, heating or explosion so as to ensure the safety of the experimenter.
26. Containers for chemical wastes shall not be left in the hallway or on the stairs, and shall not be stored in the corners of the laboratory or other invisible places.
27. When performing microbiological tests, gloves shall be worn, and after the experiment, the used glassware shall be washed through sterilization, and disposable supplies and medium shall be disposed of separately from general garbage.
28. Inflammable chemicals including acetone, oil, or gas must be kept in the place where ventilation is easy and access is infrequent.
29. Importing and exporting of high-pressure gas container must be done using transportation equipment.
30. When storing high-pressure gas containers, they must be safely and firmly fixed, isolated from flammable substances and inflammatory materials.

1. Fire Regulations.

When fire breaks out, the following guidelines shall apply.

1. Save lives.
2. Call the fire brigade.
3. Alert people in the area.
4. Extinguish the fire if possible.
5. Close doors to the area.
6. Evacuate.
7. Reassemble outside the building at the designated meeting point.

Also it is very important to know the locations of the fire-fighting equipment, fire alarms, and evacuation routes closest to the lab.
A small fire can be extinguished quickly by smothering it in a fire blanket or by spraying it with a fire extinguisher.
Lab fires in biological labs are caused most commonly by the plating of cell cultures.
To prevent this,

1. Never use paper bench coat near burners.
2. Use a low reservoir volume of ethanol.
3. Place the ethanol reservoir at least a foot from the burner and on the opposite side of the burner from the plates
4. Never wear plastic gloves when working with a flame.
5. Carry the gas container for the burner carefully with a firm grip. If it is dropped, make sure that there is no leakage by smelling for gas.
6. Notify the instructor of any gas leakage and do not light burners when there is a smell of gas.
7. Always light the match before opening the gas valve.
8. Avoid placing burners too close to overhanging shelves.
9. Never leave the table while a burner is on.

CHEMICALS

On each chemical container, there is a label that specifies the potential danger of the substance for humans and/or the environment.
Chemicals should be handled cautiously with gloves, both for your safety and for decreasing the contamination risk.
Always wear a lab coat and shoes as additional protection. Read the signs on the chemical container and the Material Safety Data Sheet
(MSDS; available online) for further direction.

BIO- SAFETY AND DISPOSAL

When you are working to generate new organisms by molecular methods, you are regulated in most countries
according to international biosafety guidelines:

1

CDC, Atlanta. Biosafety in Microbiological and Biomedical Laboratories.

2

WHO, Geneva. Laboratory Biosafety Manual, 2004.

3

ECDC, Directive 2000/54/ec of the European parliament and of the Council of 18 September 2000 on the protection of workers from risks related to exposure to biological agents at work (seventh individual directive within the meaning of Article 16(1) of Directive 89/391/EEC.

4

Institutional Bio-safety Committee, under the ministry of Health and Welfare, Korea Center for Disease Control & Prevention.

When working with microorganisms such as bacteria and viruses, there are four BioSafety Levels (BSL) numbered BSL1–4.
Biosafety level is the level of the biocontainment precautions required to isolate dangerous biological agents in an enclosed
The levels of containment range from the lowest biosafety level 1 to the highest at level 4, and Ministry of Health and Welfare and Korea Centers
for Disease Control and Prevention have specified these levels facility.
Biocontainment can be classified by the relative danger to the surrounding environment as biological
safety levels. As of 2006, there are four safety levels. These are called BSL1 through BSL4

BSL 1

1

involving well-characterized agents not known to consistently cause disease in healthy adult humans and of minimal potential hazard to laboratory personnel and the environment.

2

bacteria and viruses including canine hepatitis, non-pathogenic Escherichia coli, as well as cell cultures and non-infectious bacteria.

3

The laboratory is not necessarily separated from the general traffic patterns in the building.

4

Conducted on open bench tops using standard microbiological practices.

5

Laboratory personnel have specific training in the procedures conducted in the laboratory.

6

Supervised by a scientist with proper training.

7

Contaminated materials are left in open waste receptacles.

BSL 2

1

Similar to Biosafety Level 1 and is suitable for work involving agents of moderate potential hazard to personnel and the environment.

2

Includes various bacteria and viruses that cause only mild disease to humans, or are difficult to contract via aerosol in a lab setting such as C. difficile, most Chalmydiae, hepatitis A,B, and C, orthopoxviruses, influenza A, Lyme disease, Salmonella, mumps, measles, scapie, MRSA, and VRSA.

3

similar to Biosafety Level 1 and is suitable for work involving agents of moderate potential hazard to personnel and the environment. It includes various bacteria and viruses that cause only mild disease to humans, or are difficult to contract via aerosol in a lab setting.

4

Laboratory personnel have specific training in handling pathogenic agents,

5

Must be directed by scientists with advanced training.

6

Extreme precautions are taken with contaminated sharp items.

7

In case that infectious aerosols or splashes may be created, specified procedures should be conducted in biological safety cabinets or other physical containment equipment.

BSL 3

1

Indigenous or exotic agents which may cause serious or potentially lethal disease after inhalation.

2

Includes various bacteria, parasites and viruses that can cause severe to fatal disease in humans but for which treatments exist, such as Yersinia pestis, SARS coronavirus, Brucella, yellow fever virus.

3

Laboratory personnel have specific training in handling pathogenic and potentially lethal agents,

4

Supervised by competent scientists who are experienced in working with these agents.

5

All procedures involving the manipulation of infectious materials are conducted within biological safety cabinets, specially designed hoods, or other physical containment devices.

6

The laboratory personals should wear appropriate personal protective clothing and equipment.

7

An acceptable level of safety for the conduct of routine procedures, may be achieved in a biosafety level 2 facility, providing the filtered exhaust air from the laboratory room is discharged to the outdoors.

8

The ventilation to the laboratory is balanced to provide directional airflow into the room.

9

Access to the laboratory is restricted when work is in progress and the recommended Standard Microbiological Practices, Special Practices, and Safety Equipment for Biosafety Level 3 are rigorously followed.

BSL 4

1

Limited access only for authorized persons can work in the facilities. Not allowed for people who have weak immune system.

2

When dealing with biological hazards at this level, the use of a positive pressure personnel suit, with a segregated mandatory.

3

All works should be supervised by qualified scientists who are trained and experienced in working with these agents.

4

The entrance and exit of a level 4 bio-lab will contain multiple showers, a vacuum room, an ultraviolet light room, and other safety precautions designated to destroy all traces of the biohazard.

5

Multiple airlocks are employed and are electronically secured to prevent both doors from opening at the same time.

6

All air and water service going to and coming from a biosafety level 4 will undergo similar decontamination procedures to eliminate the possibility of an accidental release.

7

Members of the laboratory staff shall have specific and thorough training in handling extremely hazardous infectious agents, and they shall understand the primary and secondary containment functions of the standard and special practices, the containment equipment, and the laboratory design characteristics.

Conclusion

Our group’s main focus was obviously the project, but safety was an issue we always had in mind. Even when conducting experiments, safety was first ­ and we always clarified what we were doing and how we were going to do it according to our protocols before going into an experiment. We followed a rigorous code for our Biosafety Level, and conducted experiments in a safe environment.