1 biosafety

8/2/2019 1 BIOSAFETY

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Please purchase a personal

license.


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Ti liu tham kho Q s 212/2005/Q-TTg, 2005. Quy ch ca Th tng chnh phv qun l an ton sinh hc i vi cc sinh vt bin i gen.

Nguyn Vn Mi, 2008. An ton sinh hc. Nxb Gio dc.

Khut Hu Thanh v L Gia Hy, 2007. An ton sinh hc. NXB Khoahc K thut.

Barbara Eggers and Ruth Mackenzie, 2000. The Cartagena protocolon Biosafet . Oxford Universit ress.

National Research Council, 1995. Prudent practices in theLaboratory: the Handling and Disposal of Chemicals. NationalAcademy Press, Washington, D.C.

US Department of Health and Human Services, 1999. Biosafety inMicrobiological and Biomedical Laboratories. HHS publication,Washington, D.C.


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Environmental Health & Safety Lab

Safety

http://www.ehs.sunysb.edu

u e nes or esearc nvo v ngRecombinant DNA Molecules

http://www4.od.nih.gov/oba/rac/guidelines/g

uidelines.html


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Lut bo v mi trngs 55/2005-UBTVQH11 ngy 29/11/2005

Lut an ton thc phm

s 55/2010UBTVQH12 ngy 17/6/2010

Php lnh V sinh an ton thc phm

s -ngy 07/8/2003

Php lnh ging cy trng

s 15/2004/PL-UBTVQH11 ngy24/3/2004 Php lnh ging vt nui

s 16/2004/PL-UBTVQH11 ngy24/3/2004


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Quyt nh 212/2005/Q-TTg - Qui ch qun l ATSH

i vi GMO; sn phm, hng ha c ngun gc t GMO Quyt nh 79/2007/Q-TTg - K hoch hnh ng quc

n nm 2020 thc hin Cng c a dng sinh hc v

Ngh nh th Cartagena v An ton sinh hc


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SEMINAR1. Animal biosafety

2. Microorganism Biosafety (fungi, bacteria, virus)

3. Biosafety in Microbiological Laboratories

4. Biosafety in Biomedical Laboratories

5. Hazardous Chemical Waste Disposal

6. Radioactive Waste Disposal

7. Biological Waste Management

8. Manufacture and storage of hazardous materials

9. Packaging, shipping and transportation of hazardousmaterials


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10. GMOs Risk Assessment (animal, plant, food)

11. GMOs Risk Management (animal, plant, food)12. Guidelines for Biosafety

13. Cartagena on Biosafety

. , ,

15. Environmental Health & Safety Organizations

16. Biosecurity

17. Bioterrorism


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Lab SafetyLab Safety

General Safety RulesGeneral Safety Rules

1. Listen to or read instructions carefullybefore attempting to do anything.2. Wear safety goggles to protect

,materials, or things that might beable to shatter.

3. Notify your teacher if any spills or

accidents occur.


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4. After handling chemicals, always wash yourhands with soap and water.

5. During lab work, keep your hands away from

your face.6. Tie back long hair.

. .8. Know the location of the fire extinguisher, fireblanket, eyewash station, and first aid kit.

9. Keep your work area uncluttered. Take to thelab station only what is necessary.


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10. It is suggested that you wearglasses rather than contact lenses.

11. Never put anything into your mouthduring a lab experiment.

12. Clean u our lab area at the

conclusion of the laboratory period.13. Never horse around or play

practical jokes in the laboratory.


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Glassware SafetyGlassware Safety

1. Chipped or cracked glassware should not be

used. Show it to the teacher.2. Broken glassware should not be

disposed of in a classroom trashcan. There is

a spec a g ass sposa con a ner or .3. When pouring liquids into glassware,make sure the container you arepouring into is resting on a table atleast a hands breadth from the edge.


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4. Pour down a glass stirring rod to preventliquids from splattering.

5. If a piece of glassware gets broken, do nottry to clean it up by yourself. Notify theteacher.

.stopper, apply a lubricant like glycerin to theglass and use a twisting motion.

7. Do not place hot glassware in water. Rapidcooling may make it shatter.


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Chemical SafetyChemical Safety

1. Wear protective goggles and a lab

apron whenever heating or pouringhazardous chemicals.2. Never mix chemicals together unless

the manner specified).3. Never taste any chemicals (you should

never taste anything in the lab).


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4. If you need to smell the odor of a chemical,waft the fumes toward your nose with one

hand. Do not put your nose over thecontainer and inhale the fumes.

5. Never pour water into a concentrated

. .6. Follow the instructions of your teacher whendisposing of all chemicals.

7. Wash your hands after handlinghazardous chemicals.


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Electrical SafetyElectrical Safety

1. Lay electrical cords where no one can trip on

them or get caught in them.2. Be sure your hands and your lab area are dry

before using electrical equipment.

. ever po e any ng n o e ec r ca ou e s.4. Unplug cords by pulling the plug and not thecord.

5. Unplug all electrical equipment at the end ofthe lab period.


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Heating SafetyHeating Safety

1. Let burners and hotplates cool downbefore touching them. Test to see if they arecool enough by bringing the back of your handclose to them.

2. Use tongs and/or protective gloves tohandle hot objects.

3. Never reach across an open flame or

burner.


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4. The only type of glassware that maysafely be heated is either Kimax or Pyrex.

5. Always point the top ends of test tubesthat are being heated away from people.6. When heating a test tube, move it around

s ow y over e ame o s r u e e eaevenly.7. Only glassware that is thoroughly dry should

be heated.8. Heat glassware by placing it on a wire gauze

platform on a ring stand. Do not hold it in yourhand.


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9. When lighting a burner, wait until the striker is

in place before you turn on the gas.10. The amount of air can be adjusted by the air

supply valve below the tube of the burner. This

regu a es e ame empera ure an co or.11. Never leave a burner or hotplate unattended.


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First AidFirst AidInjury: Burns

Immediately flush with cold water until

burning sensation is lessened.

,

Do not touch an open wound without safetygloves. Pressing directly on minor cuts willstop bleeding in a few minutes. Apply coldcompress to bruises to reduce swelling.


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Injury: Fainting

Provide fresh air and have the personrecline so that their head is lower than therest of their body.

Flush eyes immediately with plenty of waterfor several minutes. If a foreign object islodged in the eye, do not allow the eye to

be rubbed.


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Injury: Poisoning

Find out what substance was responsible forthe poisoning and alert the teacherimmediately.

Injury: Spills on the skinFlush with large quantities of water. For

acid spills, apply baking soda solution.For base spills, apply vinegar or boric acid.

Injury: Electrical shock

Shut off the current at the source.Remove wire with rubber gloves. Alert theteacher immediately.


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Cell Lab safety

Rules of the Laboratory

Follow general lab safety guidelines from the Division of

Research Safety and the specific lab guidelines for theBioengineering Cell Culture Room

Hang coats on coat hooks behind door and keep backpacks

No eating or drinking during lab exercises or in the cellculture facility at any time

Bring safety glasses or goggles to lab when required

Always wear closed toed shoes on lab and return days Always tie back long hair and remove all jewelry

Always clean up after yourself


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Safety Procedures

- Wash hands before leaving the lab

- Label and properly store all containers- Dispose of supplies appropriately

- sharps container: pasteur pipets, glass microscopeslides razor blades needles sca els

- trash can: kimwipes, plastic serological pipets,micropipettor tips, used snap cap tubes

- Use a bleach solution to disinfect anything that

contained live cells prior to disposal in trash can

- Dispose of chemical wastes in designated waste

containers


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Invisible life forms are everywhere!

Present on skin, fall off of hair, clothing, in saliva, tears,breath.

Present on surfaces and in waterEspecially warm water

Aseptic Technique

Sterile: Devoid of all life forms (bacteria, fungi, viruses)

Difficult to achieve once people start manipulating the cultures

Aseptic: Reduced numbers of life forms

Most equipment and reagents provided to you comes sterile

gamma irradiation

autoclaving

High temperature and high pressure steam

filtration


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Aseptic Technique

work in culture hood roll back long sleeves, no rings, watches, bracelets, etc hair pulled back, caps off wash hands with antibacterial soap, spray down with 70%

ethanol

wash all surfaces with 70% EtOH dont cough, talk, sneeze, sing into hood, no eating or gum

c ew ng

lids on bottles when not in use sterile tips: change if they touch anything other than media. keep tips in hood

back surface of hood is cleanest


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Aseptic Solution Transfer Practice

Important Tips:- Loosen all caps before beginning aseptic transfer

- Keep fluids covered as much as possible

- Avoid pulling solutions up into cotton plug at top ofserological pipets

- C ange p pets etween eac u trans er

- Use smallest volume pipet possible to improve volumeaccuracy

- Pipet up and down slowly and keep pipet tip belowsolution surface when mixing to prevent bubble formation

- Avoid touching sterile eppendorf tubes not in use


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Standard Microbiological Practices

NOT permitted in laboratories:

Eating

Drinking

Smoking Handling contact lenses

P h

Storing food and drink

ALWAYS wash hands:

After handling microorganisms andanimals After removing gloves Before leaving laboratory


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Discard needles, razor blades, and scalpelblades into red, puncture resistant sharpscontainers

Dispose of broken glass into broken glass

containers, never regular trash

NEVER

recap, bend, or break needles discard needles or sharps into

biological waste bags

discard needles into regular trash


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Decontaminate all biological waste beforedisposal

Solid waste (Petri dishes, cultures): autoclaveand put in dumpster

Liquid waste: add disinfectant (bleach to10%) and pour down drain

Decontaminate work surfaces daily and afterany spill of viable material

Report accidents to the instructor

Tell Health Care Provider that you work withinfectious agents or chemicals


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CHEMICAL LAB SAFETYCHEMICAL LAB SAFETY

Chemical laboratory caution

Chemical storage

Emergency & labolatory equipments


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Chemical laboratory caution

Laboratories must place a caution sign on each

entrance door to the lab.

The NFPA diamond should contain the number of the

Place a check mark in the primary and specific hazardsections of the sign for each hazard class found in lab.

Next to the hazard symbol, list the approximate quantityof the hazard that is found in lab.


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All refrigerators, ovens, and microwaves must have LABORATORY USE

ONLY stickers attached. Household refrigerators should also have a

CAUTION DO NOT STORE FLAMMABLE SOLVENTS IN THIS BOX

sticker. The storage of flammable liquids in household refrigerators is

Radioisotope users must place a radiation hazard sticker on the doorsign next to the NFPA diamond.

Emergency numbers and information should be kept in an easily

accessible location.

Several emergency numbers are listed on every door caution sign.

Additional emergency numbers that may be needed should be

placed by the laboratory phone.

.


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Chemicals should be stored in compatible groups. Oxidizers

should never be stored with flammables, acids should never be

stored with bases, and toxics should be stored by themselves.

Chemicals from different groups that are still compatible can be

stored together.

Chemical StorageChemical Storage

Color code bottles aid in segregated chemical storage.

Liquids should also be stored away from solids.


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Hazardous chemicals (particularly corrosives) should never be

stored above the shoulder height of the shortest person in lab.

When necessary, only non-hazardous chemicals should be stored on

upper shelves.

All chemicals must be properly labeled as to the contents. Any

labels that are illegible or have fallen off should be replaced.

Secondary chemicals containers should be labeled with the contents

and the date on which the reagent was made. Labels that are loose

or illegible should be replaced. All containers must be labeled as to

their contents.


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Flammable liquids that require refrigeration must be kept in

either a flammable storage or an intrinsically safe refrigerator.They must never be stored in a household refrigerator due to

the potential for explosions.

Waste containers must be labeled as HAZARDOUS WASTE.

v

contents.

Waste containers must remain closed at all times except when waste

is being added.

Finally, hazardous waste must be placed in an appropriate containerand stored in a manner that is compatible with other wastes.


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All laboratories where flammables are stored or used must have

a fire extinguisher.

Inspects extinguishers yearly to ensure that they are ready if

needed.

Special Class D extinguishers should be kept in all labs that use

Emergency Equipment

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Inspect safety showers and eye washes yearly.

The location of each safety shower and eye wash should be

clearly posted.The area around showers and eye washes must be left

unobstructed.

Laboratory personnel should inspect eyewashes weekly.


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Fume hoods are inspected and certified yearly

Sinks should be kept clean and free of glassware. Hand washing

items such as soap and paper towels should be kept on hand.

Laboratory aisles should remain uncluttered from equipmentstorage etc.

Laboratory doors should be kept closed at all times to prevent thespread of flames should a fire occur.

Doors with magnetic closure devices may be left open while the lab

is occupied.Lab doors should remain closed when fume hoods are in operation

in order to duplicate the conditions under which the hood was

certified.


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Safety rule 11Safety rule 11-- Proper biological disposal is absolutely

essential! For cell cultures

10% sodium hypochlorite

Bleach Wait a few minutes and then rinse down sink

Bacterial and fungal cultures

AutoclaveUse autoclave bags

Cryo-worksuper cold things can injure you

Use gloves, when handling dry ice or retrieving samplesfrom a -80oC freezer or liquid nitrogen

Wear goggles when thawing samples frozen in liquid

nitrogen


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Waste managementManage waste responsible:

Classification/ Characterizing hazardous waste Proper packaging

Effective labeling

Waste collection protocol in place


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Hazardous Waste Any waste that directly or indirectly represents a threat to

human health or to the environment by introducing one ormore of the following risks:

Explosion or fire Infections, pathogens, parasites or their vectors

Chemical instability, reactions or corrosion

Acute or c ronic toxicity

Cancer, mutations or birth defects

Toxicity or damage to the ecosystems or natural resources

Accumulation in the biological food chain, persistence in

the environment or multiple effects

CHEMICAL WASTECHEMICAL WASTE


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Classification dangerous substances

Class 1 Explosives

Class 2 Gases Class 3 Flammable liquids

Class 4 Flammable solids

CHEMICAL WASTECHEMICAL WASTE

Class 5 Oxidizing substances and organic peroxides

Class 6 Toxic and infectious substances

Class 7 Radioactive Substances

Class 8 Corrosives

Class 9 Other miscellaneous substances


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Segregate chemical waste

Proper segregation =good chemical hygiene + safe workplaceenvironment

Only put compatible chemicals in a container Also do not store the following near each other

Acids and bases rgan cs an ac s

Powdered or reactive metals and combustible materialsCyanide, sulfide or arsenic compounds and acidsMercury or silver and ammonium containing compounds

Do not mix solids and liquids, Halogenated with non-halogenated chemicals


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Chemical Waste Compatibility

Incompatible chemical waste not be mixed or store together

If it must be store in same area separated secondarycontainment

Container must be compatible with the waste:

Mineral acids - Plastic

Bases -PlasticOxidizers - Glass

Organics (incl Acetic acid) - Glass


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Packaging of chemical waste

Place hazardous waste in sealable containers

Use different plastic and metal containers Sized from 25L to 200L, plastic or metal.

.

Do not leave a hazardous waste container with afunnel in it.

Glass bottles with waste must be packed withvermiculite into bigger containers.


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Packaging of chemical waste

The container should not react with the waste being stored

Similar wastes may be mixed if they are compatible Wastes from incompatible hazard classes should not be mixed

Be aware that certain metals also cause disposal problemsw en m xe w t amma e qu or ot er organ c qu s.


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Labeling

Must be labeled as HAZARDOUS WASTE

Should be accurate, legible and fully explained Contain name of the department, lab group name,

contact person details, content and concentration,azar c ass, ate

No old labels


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Storage

Select the correct container (glass / polyethylene) for storage

Use original containers if possibleUse appropriate sized container

Do not make containers too heavy to lift by the contractors

Containers must be tightly sealed and not leak

Containers correctly labeled

Container compatible with chemical being stored -separatecontainers for each type of waste

Do not store longer than 90 days


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Collection Procedures

Pack all waste in drums provided by contractors

Ensure glass bottles are prevented from breaking byadding vermiculite

All drums must be ro erl classified and labelled as

HAZARDOUS WASTE When drums are full, fill in a pick-up form

Contractors pick up the drums at your department


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Unidentified Chemical waste

Should be considered unknown hazardous waste.

Unknown waste cannot be legally transported or disposed. To dispose them safely and properly it need to be

characterised

Find out as much as possible about how the waste wasgenerated.

Please DO NOT

Pour unknown chemicals down the drain

Mix unknown chemicals with any other chemicalsBring unknown chemicals to a regular waste pick up

Abandon unknown chemicals in the work area.


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Biohazardous Waste Biohazardous waste is that waste that is capable of

producing an infectious disease in humans and

includes at a minimum blood, body fluids, discardedsharps, inoculated culture media, tissues and slides.

Biohazardous waste includes the followin cate ories:

Blood and body fluidsInfectious Sharps waste

Laboratory waste

Medical sharps

Some isolation waste

Some animal waste


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BIOLOGICAL WASTE Waste generated from biologically-cultured stocks and plates,

molecular material, blood, animal and plant tissues etc.

All sharps e.g. glass implements, needles, syringes, blades,glass Pasteur pipettes

Separate biological waste from chemical hazardous waste

Treat to eliminate biohazard by sterilization or incineration Label correct, use biohazard tape

Types of Biohazardous Waste


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Types of Biohazardous Waste

Dry Solid Contaminated Containers:

Petri Dishes

Conical Tubes

Contaminated Transfer Devices Pipette Tips

Plastic Pipettes

Liquids Pourable Wastes Stocks

Media

Blood

Aspirated Liquid Wastes

Sharps Have the ability to cut or puncture.

Pasteur pipettes

Syringes with needles

Needles

Razor Blades

Microscope slides


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Containers Biological

Biohazard/medical waste boxesDisposal of non-sharp bio hazardous waste

Cardboard box lined with a red plastic bagSeal bag with cable tie, seal box with biohaz. tape

Destruct it by high temp. incineration

Biohazard Sharp containers

Disposal sharp bio-hazardous waste

100% puncture proof

Destruction by high temp


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Waste Collection

Dry Solids (No sharps!)

Primary Containment: Collect dry, solid waste in a red bag.

The red bag must have the international biohazard symbol,the word biohazard and a label that shows address

Secondary Containment:The red bag (primary containment) must be storedin a rigid container with a lid that is resistant toleaks and punctures. The red bag must be kept inthe secondary container during use, storage, andtransport.


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Waste Collection Dry Solids

Labels containing the international biohazard symbol andthe word Biohazard have to be affixed on all 4 sides of the

secondary container and on the lid.

For secondary containers that are cylindrical, the Labels.

Sharps

Collect in a rigid puncture and leak resistant properly

labeled container. The words Biohazardous waste

Must have International Biological Hazard symbol

Name Address and Phone number


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Waste Collection Biohazardous Liquid Waste (Temporary Storage)

Collect in a non-breakable container with lid and labeled withthe international biohazard symbol and the word Biohazard.

The container needs to be in secondary containment.

Biohazardous Liquid Waste Disposal

Treat the liquid by disinfecting it with a 10% bleachsolution.

Let the solution stand for 20 minutes. Discard down sink drain, then flush with water.


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Waste Disposal Take biohazard bag and sharps container to designatedAccumulation Sites:

Life Science/Bioscience Center Receiving Area

Student Health Services Building

Generators at other locations on campus may call forbiohazard waste pick-ups

A biowaste vendor will collect biohazard bag and sharpscontainer for disposal on a weekly basis from Life Science andStudent Health Services.

Biohazardous and sharps wastes will be autoclaved while animal

carcasses will be incinerated.


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Common Mistakes in ManagingBiohazardous Waste

Containers are overfilled with waste.

Bagged waste is not tied or taped closed.

Sharps containers have pipettes sticking out, causing apotential hazard.

Biohazard label is not present on all lateral sides of thecontainer and lid.

Waste is transported to pickup container without using aproperly labeled, rigid container with tight fitting lid.

Container lids have not been cleaned.


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Other Problems

DO NOT use hazardous waste labels for biohazardouswaste.

Sharps containers must be removed seven days fromw en t ey are sea e .

Red bags must be removed seven days from initialuse or more frequently if odor poses nuisance.


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Points to remember No bio-hazardous waste shall be stored for longer

than 24 hours without being decontaminated

Decontaminated bio-hazardous waste stored upto 30 days

No storage public areas Store under refrigeration if necessary to preventodors

Sharps containers treated as regular bio-hazardous waste

i h d i k d


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Bio-hazardous waste Pick-up Procedures

Seal red bags when full with cable ties provided by

contractor Seal Biohazard/Medical waste boxes with biohazard

tape

Complete chemical/biological waste pick-up requestform

Contractors pick up waste and replace containers asneeded

Principles of Biosafety


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BIOSAFETY: Preventing lab-acquired infections

p y

Biosafety Levels

Bacteria

Viruses

Fungi

Human blood

Human cell lines Recombinant DNA

Levels of biosafety


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Levels of biosafety

BL1 - microorganisms that dont consistently causedisease in healthy adults

E. coli, S. cerevisiae, polyomavirus

Standard Microbiological Practices Basic laboratory

Unlikely to cause human disease

Sink Easily cleaned work surfaces

Impervious bench tops

Sturdy furniture Windows fitted with fly screens

Levels of biosafety


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Levels of biosafety

BL2 - microorganisms of moderate potential hazard,transmitted by contact, ingestion, puncture

Salmonella, herpesvirus, human blood

Basic laboratoryStandard Practices Plus

Trainin in handlin atho ens

Can cause disease

May be a hazard to employees

Unlikely to spread to community

Prevention or treatment available

Access to lab limitedExtreme sharps precautionsUse of Biosafety Cabinet for aerosols

BSL2 Laboratory


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BSL2 Laboratory

BSL1 Lab plus:

lockable doorschairs covered with non-fabric material

eyewash available

negative airflow, non-recirculationrecommended

door closed when work is in progress

autoclave within facility

Levels of biosafety


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f f y

BL3 - microorganisms that cause serious disease, transmittedby inhalation

M. tuberculosis, yellow fever virus, hantavirus, Y. pestis(plague)

Containment lab: double door entry; directional airflow;

BL3 Door Sign agent name

entry requirements

emergency contact(s) lab personnel

safety personnel

campus emergency responders

Levels of biosafety


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f f y

BL4 - microorganisms that cause lethal disease, withno known treatment or vaccine

Ebola virus, Marburg virus

Maximum containment lab; positive pressure ventilated

suits (moon suits)

Genetically Modified Organism (GMO)


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Genetically Modified Organism (GMO)

An organism modified by gene technology.

An organism that has inherited particular traits from anorganism (the initial organism) being traits that occurred in

the initial organism because of gene technology.

modified organism, or that belongs to a class of thingsdeclared by the regulations to be a genetically modified

organism.


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If DNA is taken from an organism, spliced to a vector, andreplaced in the same organism, then that organism is a GMO.

Just because an organism is not considered to be a GMO in

another juristiction (USA, EU) does not mean it is not a GMOin other country.

GMO does not mean that you do. You still have to apply forpermission.

Although you may not have constructed the GMO, if you

are working with it you still have to apply for permission.

Plant, animal, or fungal cells containing recombinant

DNA are considered to be GMOs.

What is not a GMO?


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What is not a GMO?

Does not include

mutants.

fusion of animal cells unless the product can form an

animal.

p ants orme x protop ast us on.

plants formed by embryo rescue or invitro fertilisation orzygote implantation.

organisms formed by natural DNA transfer.

Genetically modified organisms(GMOs) in agriculture


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(GMOs) in agriculture

Proponents argue that GMOs can offer:

Increased ag productivity & higher farm profits

Less use of chemicals

Better use of natural resources

More nutritious foods

Opponents are concerned about:

The environment

Food safety

Market power

Ethics

Three types of technical barrier to trade:


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Three types of technical barrier to trade:

1) Import bans

2) Technical standards

3) Information remedies

Standards concerning GMOs have not yet

been established but there is apossibility of import bans and a demandfor labels, which is leading to:

National GMO regulations

International trade agreements


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National GMO regulations:

European Union:

De facto moratorium since June 1999 Labelling required on all GMO-inclusive

United States:

Flexible permit procedure Labelling of GMOs is generally not required

International trade agreements:


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te at o a t ade ag ee e ts

(1) The Cartagena Protocol on Biosafety

Objective: Ensure safe transboundarymovement of GMOs

acceptGMO imports and under what conditions

Lack of scientific evidence shall not preventa country in taking action

May contain GMOs label on primaryproducts



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(2) The WTO

The WTO also acknowledges the rights of acountry to protect its environment and toensure food safety and information forco su ers.

But there are rules on how such trade-relatedmeasures may be used to achieve thesegoals.

1 biosafety

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