BIOSAFETY

CABINET
Suzette D. Itay, RMT
LEARNING
OBJECTIVES

Determine the basic principle of BSCs.

Determine the importance and functions of BSCs.

LEARNING
OBJECTIVES
Determine the start-up considerations of BSCs

Discuss the proper way to use a BSC.

Identify the do’s and don’ts when working in the BSC

CATEGORIES OF LABORATORY BIOSAFETY ACCDG.: LEVELS
BASED ON CDC
PURPOSE: FACILITATE PRECAUTIONARY MEASURES

BIOSAFETY LEVEL 1 (BSL – 1)

• Suitable for work
• Viable micro defined and with well-characterized strains
• Known not to cause dse to humans
• Most appropriate among undergrad and secondary educational training
and teaching lab
• reqs,. Basic level of containment
CATEGORIES OF LABORATORY BIOSAFETY ACCDG.: LEVELS
BASED ON CDC
PURPOSE: FACILITATE PRECAUTIONARY MEASURES

BIOSAFETY LEVEL 1 (BSL – 1)

• reqs,. Basic level of containment
• e.g. Microorganisms handled
• Bacillus subtillis
• Naegleria gruberi
• Infectiuos canine hepa virus
BIOSAFETY LEVEL 2 ( BSL – 2 )
for labs which deals with indigenous moderate – risk agents in the
community
• Containment for clinical, diagnostic and teaching labs
• Hand washing sinks
• Waste decontaminations facilities
• BSC : infectious aerosols or splashes
• Lab. Specimen
• Blood born
BIOSAFETY LEVEL 3 (BSL-3)

• Containment of indigenous or exotic agents potential for

respiratory transmission (aerosols)
• Lethal and serious infections
• Req BSC or gas-tight aerosol generation chamber
• Req controlled access to lab. Esp ventilation
• Engineering and designs
• Handling must be under supervision of experts ( scientist)
BIOSAFETY LEVEL 4 (BSL-4)
• Req. Containment when handling dangerous and exotic agents
• High individual risks of life-threatening dse.
• Transmitted via aerosol route
• No available vaccine or tx
• Class III BSC, full-body , air-supplied positive pressure suit
• Completely an isolated zone
• Adequately trained personnel
PRINCIPLE 9

•BSC is a ventilated enclosure offering

protection to the user, the product and the
environment from aerosols arising from
the handling of potentially hazardous
micro-organisms.
PRINCIPLE 10

•The continuous airflow is

discharged to the atmosphere
via a HEPA filter.
AIRFLOW 11

https://www.news-
medical.net/whitepaper/201803
20/10-Tips-to-Maximize-
Protection-When-Working-in-
Nuaire-Biological-Safety-
Cabinet.aspx
USE 12

The three States of Protection

• Personal Protection
•Product Protection
•Environmental Protection
AIRFLOW 13

https://www.news-
medical.net/whitepaper/201803
20/10-Tips-to-Maximize-
Protection-When-Working-in-
Nuaire-Biological-Safety-
Cabinet.aspx
Installation of BSCs 14

Locating BCS away from areas where

airflow patterns may be disrupted
will help protect the fragile air
curtain at the front of the cabinet.
Installation of BSCs 15

BSC should not be located directly opposite

seated work stations, other bscs, or chemical
fume hoods.

A reasonably safe distance, as determined by

a local risk assessment (LRA)
 Parts of the BSCs 17

https://microbiologynotes.org/biosafety-cabinet-
introduction-development-and-safety-guidance/
START-UP CONSIDERATIONS
18

Check that the sash is at

the appropriate height.

Check the pressure gauges

to verify that readings are
within the acceptable
range.
START-UP CONSIDERATIONS

Confirm inward airflow

-by holding a tissue at the middle of
the edge of the sash to establish
that it is drawn in.

https://www.news-medical.net/whitepaper/20180320/10-Tips-to-
Maximize-Protection-When-Working-in-Nuaire-Biological-
Safety-Cabinet.aspx
START-UP CONSIDERATIONS
• Disinfect the interior
surfaces with a disinfectant
effective against the
infectious material and
toxins used in the laboratory
START-UP CONSIDERATIONS
• Assemble all materials required for
manipulation and load into the BSC.
START-UP CONSIDERATIONS
potential for splatter or splashes: lined
with a plastic-backed absorbent pad.
Place aerosol generating
equipment towards the back of
the BSC
START-UP CONSIDERATIONS

• After loading material in the BSC,

allow sufficient time for the air to
purge and the airflow to stabilize
before initiating work.
WORKING IN THE BSC
• Perform operations as far to
the rear of the work area as
reasonable.
• Ensure that elbows and arms
do not rest on the grille or
World Health Organization, 2020
work surface.
WORKING IN THE BSC
• Avoid excessive movement of
hands and arms through the
front opening
• Arms should enter and exit
the BSC slowly and
perpendicular to the front
opening.
WORKING IN THE BSC
Keep a bottle of an appropriate
disinfectant in the BSC while work is
performed.

Segregate non- contaminated ("clean") items

from contaminated ("dirty") items.
WORKING IN THE BSC

World Health Organization, 2020

WORKING IN THE BSC
Material should be discarded in a waste container
located towards the rear of the cabinet workspace.
WORKING IN THE BSC
The work area, including the inside
surface of the window, should be
decontaminated while the BSC
remains in operation.
WORKING IN THE BSC
Natural gas and propane should not be used in
a BSC; sustained open flames

NSF International (NSF); American National Standards Institute (ANSI). (2018).

WORKING IN THE BSC

Avoid Equipment
creating air movement.
Windows that open should be kept
closed when the BSC is in use.
COMPLETION OF WORK IN THE BSC
Upon completion of work, allow
sufficient time for the air in the BSC
to purge (i.e., pass through the filter)
before disrupting the air curtain
COMPLETION OF WORK IN THE BSC
Close or cover all containers.
Disinfect the interior surfaces of the
BSC
Routinely wipe the surface of the lights
within the BSC with a suitable cleaner
or disinfectant
COMPLETION OF WORK IN THE
BSC: UV LAMPS
• Germicidal UV lamps are not substitutes
• May cause performance degradation
• May compromise personnel safety when
proper precautions are not taken
CLEANING AND DISINFECTION:

• Check electrical requirement before use.

• Turn UV lamp at least 30 minutes before use.
• For worktable and sides, use sterile or non-use
cloths
CLEANING AND DISINFECTION:

• To disinfect, use cloths damped with a

disinfecting solution
• A pre-clean to disinfecting can also be
done with water and soap.
TESTING THE BSC’S
INTEGRITY
ANNUAL TESTING

• Recommended recertification for BSC’s:

–Upon installation at customer’s site
–On-site annually in accordance with
international standards
–When the cabinet is moved
HEPA FILTER LEAK TEST (LT)

Purpose:
to check the HEPA impulsion/downflow and
exhaust filters, the filter housing, and the
mounting frames for possible leakage.
 HEPA FILTER LEAK TEST (LT)
Acceptation criteria:
– The leak (impulsion/downflow or exhaustion)
should be not more than 0.01% of the specific
before the filter

Trouble shooting: replace downflow filter/exhaust

filter
2. INFLOW VELOCITY TEST (IV)

Purpose: This test is conducted

to determine the nominal value
of the inflow velocity (air
entered to the BSC)
 2. INFLOW VELOCITY TEST (IV)

Acceptation Criteria

The nominal value is 0.45 m/s.

The average value must be within ±10% of the nominal value.

Exhaust airflow volume = 580m3/h

3. DOWNFLOW VELOCITY TEST

Purpose of the test

This test is conducted to check the nominal
value of the downflow velocity (displacement
airflow) in the work area of the safety cabinet.
3. DOWNFLOW VELOCITY TEST

Accepting criteria
The average value must be within ±10% of
the nominal value (0.36 m/s).
4. AIRFLOW PATTERN TEST

Purpose
to check the behavior of the airflows in the
sample chamber. Check to see if the
displacement airflow passes along the
entire work area
4. AIRFLOW PATTERN TEST

Accepting criteria
– The smoke shows smooth downward flow
with no dead spots or reflux. No smoke escapes
through the work aperture.
4. AIRFLOW PATTERN TEST
SUMMARY
48

• BSCs provide effective primary

containment for work with
infectious material or toxins
SUMMARY
49

• This is achieved when the BSCs are:

– Properly installed
– Properly certified or validated and
maintained
– Proper training of end-user
– Used in conjunction with good laboratory
techniques
 HEPA & ULPA FILTER
• HEPA: High Efficiency Particulate Air

• ULPA: Ultra Low Penetration Air

• Important definitions:

- HEPA: 99.99% - at 0.3 microns

- ULPA: 99.999% - at 0.12 microns

• Note: The “classical” definition of HEPA filter

is 99.97% at 0.3 microns, but nowadays all
BSC and LF in US use 99.99% at 0.3 mm
Presentation title 51

HEPA/ULPA CAPABILITY
• Removes a broad range of airborne contaminants:
– Fine dust
– Smoke
– Bacteria (typical size: 500 to 0.3 micron)
– Pollen
– Radioactive particles
– Impurity ion -> can affect Integrated Circuit speed
Presentation title 52

CLASS I BSC
• Protects personnel and
environment
• Open Front – access to
the materials
• Negative Air Pressure -
air outside of the cabinet
• HEPA filter – filter air
from the inside of the
cabinet
Presentation title
CLASS I BSC 53

- Class I BSCs are partially enclosed work stations

that protect the worker and the environment fr.
contamination.
- Class I BSCs are used in microbiology labs, in
pharmaceutical research and development, and in
cancer research labs to protect the environment
from bacteria, viruses, and carcinogens.
- The class I BSCs has an open front, for access to
the materials inside of it,
Presentation title
CLASS I BSC 54

- Negative air pressure to pull air from outside of the

cabinet,
- and a HEPA filter that air from inside the cabinet
goes through, before returning to the lab or being
vented outside, to remove the contaminants.
Presentation title 55

CLASS II BSC
• Negative-pressure ventilated cabinet
• Provides HEPA-filtered, recirculated airflow within the cabinet
• Exhaust air is HEPA-filtered
• Provides personnel and product protection
• Types of Class II BSCs
– Class II A: HEPA filtered air is discharged into the room
– Class II B: HEPA filtered air is discharged out of the room
 Presentation title 56

CLASS II TYPE A1
• 30% Exhaust, 70% Re-circulate
• Inflow velocity 75 fpm minimum
• BSL 1 –3 Usage
• Personnel and Product protection
• These cabinets do not protect against vapors and gases and, therefore, are best
used for applications requiring sterile conditions without the need to work with
volatile chemicals
• Typical uses today: Bacterial, Viral, Fungal, Parasitic
 Presentation title 57

CLASS II TYPE A2

• 30% Exhaust, 70% Re-circulate

• Inflow velocity 100 fpm minimum
• Personnel and Product protection
• Typical uses today: Bacterial, Viral, Fungal, Parasitic, Arbor-
viruses
• BSL risk level 1-3

• 70% of the airflow is recirculated within the cabinet while 30% of the air is exhausted (usually through
the top).
Presentation title 58

TISSUE CULTURE HOOD

• A class II BSCs is a partially enclosed workspace that has built in protection
for the worker, the environment, and the material inside of it.
• Class II BSCs have open fronts, a vertical laminar air flow, so outside air is run
through a filter before it gets inside, and a HEPA filter for filtering air from
inside before it goes outside the cabinet. Used in microbiology labs,
pharmaceutical labs, and cancer research labs
Presentation title 59

TISSUE CULTURE HOOD

• Class II biological safety cabinets protect the bacteria, viruses, and
carcinogens being manipulated inside the cabinet while shielding
them from outside contamination.
• There are four types of class II cabinets, A, B1, B2, and B3
•
 Presentation title 60

CLASS II TYPE B1
• 60% Exhaust, 40% Re-circulate
• Designed to protect the researcher from potentially
dangerous samples, and the samples from
contamination
• Personnel and Product protection
• Typical uses today: Bacterial, Viral, Fungal,
Parasitic, Arbor-viruses
 Presentation title
CLASS II TYPE B1 61

• 40% of air is recirculated; 60% is exhausted from the

cabinet
• Class II type B1 bscs are designed to protect the
researcher from potentially dangerous samples, and the
samples from contamination.
• These ventilated workstations are enclosed on three
sides, have a positive pressure, and are used in
microbiology labs, toxicology labs, and research labs.
Presentation title 62

CLASS II TYPE B1
• The units contain pathogens and toxic chemicals inside the work
area while at the same time protecting the samples from
environmental contamination.
• Air drawn in at a minimum inflow velocity of 100 ft/min is drawn in
through a HEPA filter and then all of the air is exhausted
through another HEPA filter.
 Presentation title 63

CLASS II TYPE B2
• 100% Exhaust

• ventilated workstations that are enclosed on three

sides, have a positive pressure

• Personnel and Product protection

• all of the air is exhausted through another HEPA filter to

the outside, as opposed to recirculating into the hood or
exhausted back into the lab

• Typical uses today: Bacterial, Viral, Fungal, Parasitic,

Arbor-viruses, Cytotoxic
Presentation title 64

BSL RISK LEVEL 1-3

- In order to protect the researcher from potentially dangerous
samples and the samples from contamination, class II type B2
BSCs are used.
- The class II type B2 BSCs are ventilated workstations that are
enclosed on three sides, have a positive pressure, and are used in
microbiology labs, toxicology labs, and research labs to contain
pathogens and toxic chemicals inside the work area while at the
same time ensuring the samples, which can include tissue and cell
cultures, are not contaminated by the environment outside of the
 Presentation title 65

CLASS III BSC

• 100% Exhaust Glove Box

• rubber gloves that are attached to the cabinet

• Outside air is filtered through a HEPA filter prior to

entry, and air leaving the class III biological safety
cabinet is filtered through two HEPA filters before
being vented outside

• Anaerobic chambers are also available

• Typical uses today: Toxic Powders, BSL 4 Agents

Presentation title 66

- Class III BSCS, also known as glove boxes or isolation glove

boxes, are enclosed work areas designed to protect the worker,
the environment, and the sample from contamination.
- The laboratory technician can manipulate materials inside the
class III BSCs by using rubber gloves that are attached to the
cabinet.
Presentation title 67

- Outside air is filtered through a HEPA filter prior to entry, and air leaving the
class III BSC is filtered through two HEPA filters before being vented outside.
- Anaerobic chambers are also available. These units allow researchers to
easily process, culture and examine samples without exposure to atmospheric
oxygen.
- Accessories for Class III cabinets include heating elements, filter and
ionizers.
Presentation title 68

SELECTION OF A SAFETY CABINET THROUGH RISK ASSESSMENT

Presentation title 69

POORLY-LOCATED BSCS
Presentation title 70

WELL-LOCATED BSCS
THANK YOU

References

NSF International (NSF); American National Standards Institute (ANSI). (2018).

Biosafety Cabinetry: Design, Construction, Performance, and Field Certification
(NSF/ANSI Standard 49-2018) Retrieved from https://ehs.unl.edu/sop/s-bio-
cabinet.PDF

World Health Organization.(2020). Laboratory Biosafety Manual; Biological safety

cabinets and other primary containment devices.. 4th Edition Retrieved from
file:///C:/Users/Suzette%20D.%20Itay/Downloads/9789240011311-eng.pdf