11

Presented by:

UCLA EH&S BioSafety Program

Safe Use of Biological Safety

Cabinets

2

Objectives

Briefly discuss Chemical Fume Hoods & Laminar Flow Cabinets

Discuss how Biological Safety Cabinets (BSC) function

Review general guidelines for work preparations

Discuss general cabinet procedures

Discuss general clean up operations

Show a video on improvement of work habits

Take & Review Quiz

2

33

Chemical Fume Hood

Fume Hoods are

a type of local ventilation device that is designed to limit

inhalation exposure to hazardous or noxious fumes, vapors

or dusts.

Air is drawn in from the front (open) side of the hood and

expelled outside the building

There are different types of fume hoods which serve a

particular purpose and are installed depending upon the labs

use.

(e.g. traditional fume hoods (Constant Air Volume (CAV) or

Variable Air Volume (VAV)), bench top fume hoods, and

ductless fume hoods.)

Contact Lab Safety for further information at:

laboratorysafety@ehs.ucla.edu44

Personal?

Product?

Lab Environment?

YES

NO

YES, when ducted

What types of protection does a

Chemical Fume Hood Offer?

55

Chemical Fume Hoods

Are NOT Biological Safety Cabinets

Do not use anything pathogenic

Designed to protect the worker by capturing chemical fumes, vapors

and dusts and sweeping them away from the worker

Does not protect the product from contaminants

Various types exist so contact Lab Safety before beginning work in

any Fume Hood

Must be certified Annually

To schedule email: fumehoods@ehs.ucla.edu

Typically Used for:

Working with chemicals

Trace amounts of radiation

6

A Laminar Flow Cabinet is(NOT A BSC)

a carefully enclosed bench designed to provide protection for your products

and samples from aerosols and contaminants present in ambient air where

operator protection is not required.

Air is drawn through a HEPA filter, blown in a laminar flow in either a

horizontal or vertical pattern.

It is usually made of stainless steel with no gaps or joints where spores

might collect.

May have a UV-C germicidal lamp to sterilize the shell and contents when

not in use.*

*It is important to switch UV-C lights off during use, they can cause damage to the skin & eyes.

Laminar Flow Cabinet

Clean Bench

77

Personal?

Product?

Lab Environment?

NO

YES

NO

What types of protection does a

Laminar Flow Cabinet Offer?

88

Horizontal Laminar Flow Bench

Are NOT Biological Safety Cabinets

Do NOT use with infectious or hazardous agents!!!

Offers product protection ONLY

Will NOT protect the operator!!!

Typically used for:

Preparation of intravenous mixtures

Plant tissue culture

Electronic and mechanical assembly

Pharmaceutical procedures

Media preparation

99

3 Types of Protection are:

Personnel Product Lab Environmental

Biological Safety Cabinets (BSC) areenclosed, ventilated workspaces for safely working with materials

contaminated with (or potentially contaminated with) pathogens in a lab.

BSCs are a type of engineering control for minimizing air contamination

Several different types exist, differentiated by the type of protection provided

Biological Safety Cabinets (BSC)

10

HEPA Filters

HIGH

EFFICIENCY

PARTICULATE

AIR

Purpose of a HEPA filter is to

Capture airborne particles and create a particle free work environment

(e.g. clean rooms, clean benches)

1111

HEPA Filters

If particle size = 0.3 micron then the filter efficiency is 99.97%.

If particle size is < 0.3 microns; or > 0.3 microns the filter efficiency increases.

Does not filter vapor or gas12

BioSafety Cabinets (BSC)

The U.S. Centers for Disease Control and Prevention (CDC) classifies

BSCs into three classes:

Class I Personnel & lab environment protection, no product protection

Class II Personnel, lab environment & product protection

Class III maximum personnel & lab environment protection, product protection

These classes are distinguished in two ways:

1. the level of personnel & lab environment protection provided

2. the level of product protection provided

12

1313

Biosafety Cabinets Class I

Class I

provide personnel and environmental protection but no product protection

Uses HEPA filters to remove airborne particles

Inward airflow (similar to a fume hood) can cause contamination of samples

Inward airflow is maintained at a minimum velocity of 75 ft/min.

Common uses:

to enclose specific equipment (e.g. centrifuges); or

procedures that potentially generate aerosols (e.g. animal cage changing)

Exhaust can be

Ducted - to the building exhaust system; or

Un-Ducted - recirculating HEPA filtered exhaust back into the lab environment

1414

Biosafety Cabinets Class II

Class II

provide personnel, lab environment & product protection

Uses HEPA filters to remove airborne particles

Uses a Laminar flow to reduce air turbulence and aids in capture of

airborne contaminants

There are four Types of Class II BSCs: A1, A2, B1, B2

Exhaust method varies by Type

Common uses:

Work with sterile animal tissue and cell culture systems

Propagation of viruses

1515

Biosafety Cabinets - Class II

Class II A2 Class II B1Class II A1 Class II B2

Be cautious of the air curtain when using!

Do not block the vents

Do not use sweeping or rapid arm movements

Minimize movement of materials in and out of the cabinet

Disrupting the air curtain increases the risk for contaminant of your product or

release into the lab

_

1616

Biosafety Cabinets - Class II

Type A1

Intake

Minimum

75 FPM

Exhaust HEPA filtered air may be recirculated into

room or exhausted outdoors

Have HEPA filtered recirculated air mixed with

HEPA filtered inflow air

70% of air is recirculated

May have positive pressure contaminated ducts

and plenums

NO volatile toxic chemicals or volatile

radionuclides

Chemical vapors can build up in the cabinet and

become a health & safety hazard

17

Biosafety Cabinets - Class II

Type A2

Exhaust HEPA filtered air may be

recirculated into room or directed outdoors

Have HEPA filtered recirculated air mixed

with HEPA filtered inflow air

70% of air is recirculated

All biologically contaminated ducts and

plenums under negative pressure

Generally, no volatile toxic chemicals or

volatile radionuclides

Chemical vapors can build up in the cabinet

and become a health & safety hazard

Intake

Minimum

100 FPM

1818

Biosafety Cabinets - Class II

Type B1

Intake

Minimum

100 FPM

HEPA filtered down flow air composed largely of uncontaminated recirculated inflow

30% of air is recirculated

Split work surface

Front of work surface = filtered and recirculated

Back of work surface = filtered and exhausted through building exhaust

Building exhaust system must pull air out of the BSC

Must be Hard-Ducted

Have all biologically contaminated ducts and plenums under negative pressure

Can use with minute quantities of volatile toxic chemicals and trace amounts of radionuclides (contact EHS first)

_

1919

Biosafety Cabinets - Class II

Type B2

Intake

Minimum

100 FPM

Have HEPA filtered down flow air drawn from

the laboratory or the outside

No recirculation

Have all contaminated ducts and plenums

under negative pressure or surrounded by

negative pressure ducts & plenums

Exhaust circulating air to the outside through

building exhaust after HEPA filtration.

MUST be Hard-Ducted to dedicated exhaust

Can use volatile toxic chemicals and

radionuclides (contact EHS first)

2020

Biosafety Cabinet Location

Keep away from all other sources of airflow

Open doors & windows

Personnel traffic

Supply air diffuser

Other lab equipment

Personal & Product protection are compromised as cross drafts approach air intake!

Must be seismically stabilized

(i.e. bolted to the floor, anchored to the building etc.)

2121

Maximum Personal & Lab Environment protection and some Product

Protection.

Often customized for specific materials and procedures.

Commonly used for work with:

Highly infectious microbiological agents

Conducting hazardous operations

BSL 4 Laboratories

Biosafety Cabinets - Class III

2222

What Do BSCs Have In Common?

Continuous inward airflow protects you from aerosols

Exhaust air is HEPA filtered to protect the lab environment from becoming contaminated with aerosols

HEPA-filtered downward laminar airflow over the workspace protects the product from contamination (excluding Class I)

2323

UV-C Light in BSC

UCLA Does Not recommend the use of UV-C light to sterilize biosafety cabinets.

Can cause skin cancer and burn the cornea!

Without proper maintenance the CDC does not recommend using UV-C light Must be cleaned regularly to keep clear of dust

Must be monitored to ensure that the appropriate intensity of UV-C light is emitted (254 nm)

Shadows? Dont get touched by the UV-C rays!

No one is allowed in the room while the UV-C light is on.

2424

BSC must be certified when:

1. New (before first use)

2. Every year

3. If repaired

4. If relocated

(even if moved within the same room)

UCLA has a contract with a company called TSS for BSC certification

2525

Guidelines for working in the BSC

Never

eat, drink, chew gum, store food, or smoke nearby

2626

Guidelines for Working in the BSC

Schedule uninterrupted work times

Store papers or pencils outside of the BSC

One person working in a BSC at a time

Have arm pits level with bottom of the window sash

Use slow movements in BSC

Minimize entering and exiting the BSC

Do not block any of the grills!!

Do not store equipment or supplies in or on the BSC

2727

Guidelines for Work Preparations

1. Check certification before using!

2. Let BSC run for at least 3 min before using

3. Disinfect all surfaces of the BSC

4. Properly position the window sash

5. Close drain valve (if there is one)

6. Make sure that all materials needed are inside the BSC

7. Segregate clean items from ones that will get contaminated

2828

Guidelines for Working in the BSC

UCLA POLICY No open flames in the BSC!*

Why?

Disrupts airflow

Excessive heat builds up

Possible gas build up

Damages HEPA Filter

*Alternatives do exist. Contact the BioSafety department for more information.

2929

Dont let this be your Biosafety Cabinet!

3030

BSC Cleanup

When do you clean your BSC?

Before and after every procedure!

What do you use to clean your BSC?

Alcohol (70% solution)

Bleach (10% solution)

Approved Disinfectant Agent

Keep in mind that bleach can be corrosive!

3131

Guidelines for BSC Cleanup

1. Place all items that may have come in contact with the agent(s), such as the used pipettes, in an appropriate biohazard or other waste container.

2. Disinfect surfaces of all equipment used.

3. Remove all items from inside the BSC

4. Disinfect all interior surfaces of the BSC

* If possible leave the BSC running

3232

BSC Cleanup

If the BSC must be turned off:

1. Do so after a final purge of at least 3 minutes

2. Close the cabinet window sash completely

3333

Validation of Decontamination

Use RODAC plates to detect presence of microorganisms on surfaces

Fill with media appropriate for agents you are using

Incubate for 24 hours and check for growth

3434

Technical Safety Services (TSS)

To schedule service with TSS call

1-800-877-7742

Los Angeles Office

1-562-694-3626

TSS should schedule service in 1 to 5 days

Usually within 48 hours

3535

Contact Biosafety for consultation or questions:

Email: biosafety@ehs.ucla.edu

Website: www.biosafety.ucla.edu

Main Line: (310) 206-3929

For a copy of your EH&S Training History email:

training@ehs.ucla.edu

Envi

ronm

ent,

Hea

lth a

nd S

afet

y

Page

1 o

f 1

Bio

safe

ty P

rogr

am, I

BC

Effe

ctiv

e D

ate

Sept

embe

r 25,

200

8

Ope

n Fl

ames

in B

iolo

gica

l Saf

ety

Cab

inet

s IB

C P

olic

y

Polic

y on

the

Use

of O

pen

Flam

es in

Bio

logi

cal S

afet

y C

abin

ets

In

trod

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n In

acc

orda

nce

with

the

CD

C B

iosa

fety

in M

icro

biol

ogic

al a

nd B

iom

edic

al L

abor

ator

ies (

5th e

d.):

O

pen

flam

es a

re n

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quire

d in

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near

mic

robe

-fre

e en

viro

nmen

t of a

bio

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cal s

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y ca

bine

t. O

n an

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the

neck

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cul

ture

ves

sel w

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hich

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ng in

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e tu

be o

r fla

sk. A

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en fl

ame

in a

BSC

, how

ever

, cre

ates

turb

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rupt

s the

pa

ttern

of H

EPA

-filt

ered

air

bein

g su

pplie

d to

the

wor

k su

rfac

e. W

hen

deem

ed a

bsol

utel

y ne

cess

ary,

touc

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d w

ith a

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ay b

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ir di

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bur

ner m

ust b

e tu

rned

off

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ork

is c

ompl

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. Sm

all e

lect

ric "

furn

aces

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ava

ilabl

e fo

r dec

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min

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ps a

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eedl

es a

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re

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(WH

O) L

abor

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y Bi

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Man

ual:

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es sh

ould

be

avoi

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in th

e ne

ar m

icro

be-f

ree

envi

ronm

ent c

reat

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the

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w p

atte

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are

als

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o st

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oops

, mic

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rner

s or e

lect

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ar

e av

aila

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are

pref

erab

le to

ope

n fla

mes

.2

In li

ght o

f rec

ent e

vent

s at U

CLA

invo

lvin

g fir

es in

side

of b

iosa

fety

cab

inet

s, th

e In

stitu

tiona

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safe

ty

Com

mitt

ee (I

BC

) has

ado

pted

a g

uida

nce

polic

y fo

r the

use

of o

pen

flam

es in

side

of b

iosa

fety

cab

inet

s. Po

licy

In

con

side

ratio

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afor

emen

tione

d C

DC

and

WH

O g

uide

lines

, rec

omm

enda

tions

from

the

man

ufac

ture

rs

of b

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fety

cab

inet

s, la

bora

tory

safe

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sk a

sses

smen

ts, r

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t inc

iden

ts in

volv

ing

fires

insi

de o

f bio

safe

ty

cabi

nets

, and

oth

er fa

ctor

s, th

e U

CLA

IBC

has

ado

pted

the

follo

win

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idan

ce p

olic

y:

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IBC

stro

ngly

reco

mm

ends

that

the

prac

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sing

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nd B

unse

n bu

rner

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s tha

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r be

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cont

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med

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rnat

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B

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e re

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datio

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r saf

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tern

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usi

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abin

et.

If y

our l

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ific

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he u

se o

f an

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of a

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ty c

abin

et p

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e co

nsul

t w

ith th

e IB

C.

Alte

rnat

ives

incl

ude:

Flam

e-on

-dem

and

Bur

ners

Elec

tric

burn

ers

D

ispo

sabl

e to

ols

For m

ore

info

rmat

ion

abou

t the

se sa

fety

gui

delin

es, r

esou

rces

, saf

e al

tern

ativ

es to

Bun

sen

burn

ers,

bios

afet

y ca

bine

t tra

inin

g, o

r abo

ut th

is p

olic

y, c

onta

ct th

e U

CLA

Off

ice

of E

nviro

nmen

t, H

ealth

& S

afet

y B

iosa

fety

at

310-

206-

3929

, em

ail b

iosa

fety

@eh

s.ucl

a.ed

u or

vis

it ht

tp://

ww

w.b

iosa

fety

.ucl

a.ed

u/.

Ref

eren

ces

1.

Bio

safe

ty in

Mic

robi

olog

ical

and

Bio

med

ical

Lab

orat

orie

s, 5t

h Ed

ition

, App

endi

x A

: Prim

ary

Con

tain

men

t for

Bio

haza

rds:

Sel

ectio

n, In

stal

latio

n an

d U

se o

f Bio

logi

cal S

afet

y C

abin

ets,

UR

L:

http

://w

ww

.cdc

.gov

/od/

ohs/

bios

fty/b

mbl

5/bm

bl5t

oc.h

tm

2.

Wor

ld H

ealth

Org

aniz

atio

n La

bora

tory

Bio

safe

ty M

anua

l, 3r

d Edi

tion,

Par

t III

: Lab

orat

ory

Equi

pmen

t, U

RL:

http

://w

ww

.who

.int/c

sr/d

elib

epid

emic

s/W

HO

_CD

S_C

SR_L

YO

_200

4_11

/en/

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