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Environment, Health, & Safety Division

 

Appendix E

Biosafety Cabinets

 

E.1    Introduction and Scope

Biological safety cabinets or biosafety cabinets (BSCs) are hoods with high-efficiency particulate air (HEPA) filters that provide personnel, environmental, and product protection when appropriate practices and procedures are followed. Key BSC information and requirements are summarized in Section 5.6.4.2. This appendix provides the following information and requirements on BSCs:

  • Classifications
  • Work practices and procedures
  • Decontamination
  • Installation and engineering
  • Testing and certification

Information in this appendix primarily contains information that was excerpted and adapted from Appendix A (Primary Containment for Biohazards:  Selection, Installation, and Use of Biological Safety Cabinets) of Biosafety in Microbiological and Biomedical Laboratories (BMBL), fifth edition, and minimally reiterates LBNL policies presented in Section 5.6.4.2 (Hoods and Biosafety Cabinets) of this manual.

E.2    Biosafety Cabinet Classifications

Three primary types of BSCs have been developed to meet varying research and clinical needs. These primary BSC types are designated as Class I, II, and III. Class II BSCs are also further subdivided into different Class II types. Tables E-1 and E-2 summarize the similarities and differences in the types of protection and physical characteristics of different classes of BSCs. The sections following these tables summarize and illustrate the characteristics of BSC classes used at LBNL. This information should be used in BSC selection and risk assessment.

Table E-1
Protection Offered by Classes of Biosafety Cabinets

Biological
Risk Assessed

Protection Provided

 

Personnel

Product

Environmental

BSC Class

Biosafety Level (BL) 1 to 3

Yes

No

Yes

I

BL1 to 3

Yes

Yes

Yes

II (A1, A2, B1, B2)

BL4

Yes

Yes

Yes

III
II - when used in room with suit

Source: adapted from BMBL, fifth edition, Appendix A, Table 1.


Table E-2
Characteristics of Biosafety Cabinet Classes

 

BSC Class, Type

 

Face Velocity
(fpm)

 

Airflow Pattern

Applications

Nonvolatile Toxic Chemicals and
Radionuclides

Volatile Toxic Chemicals and  Radionuclides

I

75

In at front through HEPA to the outside or into the room through HEPA (Figure 1)

Yes

When exhausted
outdoors 1,2

II, A1

75

70% recirculated to the cabinet work area through HEPA; 30% balance can be exhausted through HEPA back into the room or to outside through a canopy unit

Yes
(minute amounts)

No

II, A2

100

Similar to II, A1, but has 100 linear fpm intake air velocity and plenums are under negative pressure to room (Figure 2); exhaust air can be ducted to outside through a canopy unit (Figure 3)

Yes

When exhausted
outdoors
(formerly "B3")
(minute amounts) 1,2

II, B1

100

30% recirculated, 70% exhausted. Exhaust cabinet air must pass through a dedicated duct to the outside through a HEPA filter (Figure 4)

Yes

Yes
(minute amounts)1,2

II, B2

100

No recirculation; total exhaust to the outside through a HEPA filter

 

Yes

Yes. (small amounts) 1,2

III

N/A

Supply air is HEPA filtered. Exhaust air passes through two HEPA filters in series and is exhausted to the
outside via a hard connection (Figure 5)

Yes

Yes
(small amounts) 1,2

 

Footnotes:

  • Installation may require a special duct to the outside, an in-line charcoal filter, and a spark-proof (explosion-proof) motor and other electrical components in the cabinet. Discharge of a Class I or Class II Type A2 cabinet into a room should not occur if volatile chemicals are used.
  • In no instance should the chemical concentration approach the lower explosion limits of the compounds.

Source: adapted from BMBL, fifth edition, Appendix A, Table 2.

E.2.1    Class I Biosafety Cabinet

The Class I BSC provides personnel and environmental protection, but no product protection. It is similar in air movement to a chemical fume hood, but has a HEPA filter in the exhaust system to protect the environment. Figure 1 shows a diagram of a Class I BSC.

Figure 1. Class I BSC. (A) front opening, (B) sash, (C) exhaust HEPA filter, and (D) exhaust plenum. Note: The cabinet needs to be hard connected to the building exhaust system if toxic vapors are to be used. Source: BMBL, fifth edition, Appendix A.

E.2.2    Class II Biosafety Cabinet

Class II BSCs (Types A1, A2, B1 and B2) provide personnel, environmental, and product protection. Airflow is drawn into the front grille of the cabinet, providing personnel protection. In addition, the downward laminar flow of HEPA-filtered air provides product protection by minimizing the chance of cross-contamination across the work surface of the cabinet. Because cabinet exhaust air is passed through a certified HEPA filter, the exhaust air is particulate-free (environmental protection), and may be recirculated to the laboratory (i.e., Type A1 and A2 BSCs only) or discharged from the building via the exhaust duct system and a canopy connection. Exhaust air from Types B1 and B2 BSCs must be discharged to the outdoors via a hard duct connection. Figure 2 shows a diagram of a Class II Type A2 BSC, which is the most common type of BSC at LBNL. Figure 3 shows a diagram of a canopy (or thimble) unit that is normally required when connecting a Class II Type A1 or A2 BSC to an exhaust duct system. Figure 4 shows a Class II, Type B1 BSC, which is also used at LBNL. Installation of a Class II, Type B1 BSC typically requires a hard duct connection to the exhaust system without a canopy or thimble unit connection.

HEPA filters are effective at trapping particulates and thus infectious agents but do not capture volatile chemicals or gases. Only Type A2 exhausted or Types B1and B2 BSCs exhausting to the outside should be used when working with volatile toxic chemicals, but amounts must be limited. See Table 2 for additional information.



Figure 2.
Class II, Type A2 BSC. Tabletop model. (A) front opening, (B) sash, (C) exhaust HEPA filter, (D) supply HEPA filter, (E) positive-pressure common plenum, (F) negative-pressure plenum. Unless it is connected to the building exhaust system, the Class II, Type A2 BSC is not equivalent to what was formerly called a Class II, Type B3 BSC. Note: The Class II, Type A2 BSC should be canopy connected to the exhaust system. Source: adapted from BMBL, fifth edition, Appendix A.



Figure 3.
Canopy (Thimble) Unit. Canopy (thimble) units for connecting a Class II, Type A1 or A2 BSC to the exhaust duct system. (A) balancing damper, (B) flexible connector to exhaust system, (C) cabinet exhaust HEPA filter housing, (D) canopy unit, (E) BSC. Note: There is a one-inch gap between (D) the canopy unit and (E) the exhaust filter housing through which room air is exhausted. Source: adapted from BMBL, fifth edition, Appendix A.



Figure 4.
Class II, Type B1 BSC (classic design). (A) Front opening, (B) sash, (C) exhaust HEPA filter, (D) supply HEPA filter, (E) negative-pressure dedicated exhaust plenum, (F) blower, (G) additional HEPA filter for supply air. Note: The cabinet exhaust needs to be hard connected to the building exhaust system. Source: BMBL, fifth edition, Appendix A.

E.2.3    Class III Biosafety Cabinet

A standard Class III BSC (Figure 5) is designed for working with highly infectious microbiological agents and conducting hazardous operations. It is a gas-tight enclosure with a nonopening view window that provides maximum protection for the environment and the worker. Access for passage of materials into the cabinet is through a chemical dunk tank accessible through the cabinet floor, or a double-door pass-through box (e.g., an autoclave) that can be decontaminated between uses. Reversing that process allows materials to be removed from the Class III BSC safely. Both supply and exhaust air pass through a HEPA filter on a Class III cabinet. Exhaust air must pass through two HEPA filters, or a HEPA filter and an air incinerator, before discharge to the outdoors. Airflow is maintained by a dedicated, independent exhaust system exterior to the cabinet, which keeps the cabinet under negative pressure (minimum pressure of 0.5 inches of water gauge). Some Class III BSCs may not have all of these controls, based on the risk assessment conducted (e.g., types of materials and manner of work).

Long, heavy-duty rubber gloves are attached in a gas-tight manner to ports in the cabinet and allow direct manipulation of the materials isolated inside and prevent the user's direct contact with the hazardous materials. Depending on the design of the cabinet, the supply HEPA filter provides particulate-free, albeit somewhat turbulent, airflow within the work environment. Laminar airflow is not a characteristic of a Class III cabinet.

 

Figure 5.
Class III BSC. (A) Glove ports with O-ring for attaching arm-length gloves to cabinet, (B) sash, (C) exhaust HEPA filter, (D) supply HEPA filter, (E) double-ended autoclave or pass-through box. The cabinet exhaust needs to be hard connected to an independent dedicated exhaust system. The exhaust air must be double HEPA filtered or HEPA filtered and incinerated. Source: adapted from BMBL, fifth edition, Appendix A.

E.2.4    Clean Benches (Not BSCs)

Horizontal and vertical laminar flow “clean benches” are shown in Figures 6 and 7. These units may provide protection for the product, but are not considered safety hoods or BSCs and must not be used for infectious or toxic materials or when a hood or BSC is needed to protect the worker.



Figure 6.
Horizontal Laminar Flow “Clean Bench.” (A) Front opening, (B) grille, (C) supply HEPA filter, (D) plenum, (E) blower, (F) grille. Source: BMBL, fifth edition, Appendix A.




Figure 7.
Vertical Laminar Flow “Clean Bench.” (A) Front opening, (B) sash, (C) supply HEPA filter, (D) blower. Source: BMBL, fifth edition, Appendix A.

E.3    Biosafety Cabinet Work Practices and Procedures

This section discusses in detail standard work practices and procedures for investigators working in a Class II BSC. In general, these practices and procedures are important for protection of the worker or the product, but the importance of each practice or procedure for the safety of the worker often depends on the nature of biological materials and the work being conducted. A shorter list of key BSC work practices and procedures is provided in Appendix D.

E.3.1    Preparing for BSC Work

This section discusses preparing for work within a Class II BSC.

Air Current Disruptions. Preparing a written checklist of materials necessary for a particular activity and placing necessary materials in the BSC before beginning work minimizes the number and extent of air curtain disruptions compromising the fragile air barrier of the cabinet. The rapid movement of a worker's arms in a sweeping motion into and out of the cabinet will disrupt the air curtain and compromise the partial containment barrier provided by the BSC. Moving arms slowly in and out and perpendicular to the face while opening the cabinet will reduce this risk. Other personnel activities in the room (e.g., rapid movements near the face of the cabinet, walking traffic, room fans, open/closing room doors, etc.) may also disrupt the cabinet air barrier.

Personal Protective Equipment (PPE). Eye protection and laboratory coats buttoned over street clothing must be worn. Latex, vinyl, nitrile, or other suitable gloves must be worn to provide hand protection. Higher levels of PPE can be included as determined by an individual risk assessment. For example, a solid front, back-closing laboratory gown provides better protection of personal clothing than a traditional laboratory coat and is a recommended practice at BL3.

Body and Material Positioning. Before beginning work, the BSC user should adjust the stool height so that his/her face is above the front opening. Manipulation of materials should be delayed for approximately one minute after placing the hands/arms inside the cabinet. This allows the cabinet to stabilize, the user to "air sweep" his or her hands and arms, and to allow time for turbulence reduction. When the user's arms rest flatly across the front grille, the arms may occlude the grille opening, and room air laden with particles may flow directly into the work area rather than being drawn down through the front grille. Raising the arms slightly will alleviate this problem. The front grille must not be blocked by towels, research notes, discarded plastic wrappers, pipetting devices, etc. All operations should be performed on the work surface at least four inches from the front grille. If there is a drain valve under the work surface, it should be closed prior to beginning work in the BSC.

Materials or equipment placed inside the cabinet may cause disruption of the airflow, resulting in turbulence, possible cross-contamination and/or breach of containment. Extra supplies (e.g., additional gloves, culture plates or flasks, culture media) should be stored outside the cabinet. Only the materials and equipment required for immediate work should be placed in the BSC.

Purge and Decontamination. If the cabinet has been shut down, the blowers should be operated at least four minutes before beginning work to allow the cabinet to "purge." This purge will remove any suspended particulates in the cabinet. The work surface, the interior walls (except the supply filter diffuser), and the interior surface of the window should be wiped with 70% ethanol (EtOH), a 1:100 dilution of household bleach (i.e., 0.05% sodium hypochlorite), or other disinfectant as determined by the investigator to meet the requirements of the particular activity. When bleach is used, a second wiping with sterile water is needed to remove the residual chlorine, which may eventually corrode stainless steel surfaces. Wiping with nonsterile water may recontaminate cabinet surfaces, a critical issue when sterility is essential (e.g., maintenance of cell cultures).

Similarly, the surfaces of all materials and containers placed into the cabinet should be wiped with 70% EtOH to reduce the introduction of contaminants to the cabinet environment. This simple step will reduce the introduction of mold spores and thereby minimize contamination of cultures. The further reduction of microbial load on materials to be placed or used in BSCs may be achieved by periodic decontamination of incubators and refrigerators.

E.3.2    Material Placement inside the BSC

This section covers placement of materials inside the BSC.

Surface Towels. Plastic-backed absorbent towels can be placed on the work surface but not on the front or rear grille openings. The use of towels facilitates routine cleanup and reduces splatter and aerosol generation during an overt spill. It can be folded and placed in a biohazard bag or other appropriate receptacle when work is completed.

Inside Materials and Sash. All materials should be placed as far back in the cabinet as practical, toward the rear edge of the work surface and away from the front grille of the cabinet (Figure 8). Similarly, aerosol generating equipment (e.g., vortex mixers, tabletop centrifuges) should be placed toward the rear of the cabinet to take advantage of the split in downward laminar air flow to the front and rear grilles as the air approaches the work surface. Bulky items such as biohazard bags, discard pipette trays, and vacuum collection flasks should be placed to one side of the interior of the cabinet. If placing those items in the cabinet requires opening the sash, make sure that the sash is returned to its original position before work is initiated. The correct sash position (usually 8 or 10 inches above the base of the opening) should be indicated on the front of the cabinet. On most BSCs, an audible alarm will sound if the sash is in the wrong position while the fan is operating.

Practices Do and Do Not Interfere with BSC Operation. Certain common practices interfere with the operation of the BSC. The biohazard collection bag should not be taped to the outside of the cabinet. Upright pipette collection containers should not be used in BSCs nor placed on the floor outside the cabinet. The frequent inward/outward movement needed to place objects in these containers is disruptive to the integrity of the cabinet air barrier and can compromise both personnel and product protection. Only horizontal pipette discard trays containing an appropriate chemical disinfectant should be used within the cabinet. Furthermore, potentially contaminated materials should not be brought out of the cabinet until they have been surface decontaminated. Alternatively, contaminated materials can be placed into a closable container for transfer to an incubator, autoclave, or another part of the laboratory.

E.3.3    Operations within a Class II BSC

Splatters and Aerosols. Many procedures conducted in BSCs may create splatter or aerosols. Good microbiological techniques should always be used when working in a BSC. For example, techniques used to reduce splatter and aerosol generation will also minimize the potential for personnel exposure to infectious materials manipulated within the cabinet. Class II cabinets are designed so that horizontally nebulized spores introduced into the cabinet will be captured by the downward flowing cabinet air within 14 inches of travel. Therefore, as a general rule of thumb, keeping clean materials at least one foot away from aerosol-generating activities will minimize the potential for cross-contamination.

Work Flow. The work flow should be from "clean to dirty" (see Figure 8). Materials and supplies should be placed in the cabinet in such a way as to limit the movement of "dirty" items over "clean" ones. Several measures can be taken to reduce the chance for cross-contamination of materials when working in a BSC. Opened tubes or bottles should not be held in a vertical position. Investigators working with petri dishes and tissue culture plates should hold the lid above the open sterile surface to minimize direct impaction of downward air. Bottle or tube caps should not be placed on the towels. Items should be recapped or covered as soon as possible.


Figure 8. Typical Work Layout Inside a BSC. Shown above is a typical layout for working “clean to dirty” within a Class II BSC. Clean cultures (left) can be inoculated (center); contaminated pipettes can be discarded in the shallow pan, and other contaminated materials can be placed in the biohazard bag (right). This arrangement is reversed for left-handed persons. Source: adapted from BMBL, fifth edition, Appendix A.

Burners and Open Flames. Open flames are not required in the near-microbe-free environment of a BSC. On an open bench, flaming the neck of a culture vessel will create an upward air current, which prevents microorganisms from falling into the tube or flask. An open flame in a BSC, however, creates turbulence that disrupts the pattern of HEPA-filtered air being supplied to the work surface and may cause fires. When deemed absolutely necessary, touch-plate microburners equipped with a pilot light to provide a flame on demand should be used. These burners will minimize internal cabinet air disturbance, heat buildup, and fire risk. The burner must be turned off when work is completed. Small electric "furnaces" are also available for decontaminating bacteriological loops and needles, and are preferable to an open flame inside the BSC. Disposable or recyclable sterile loops should be used whenever possible.

Text Box:    Text Box:
     

A fire inside a BSC occurred when the gas rubber hose connected to a Touch-O-Matic Bunsen burner melted and gas in the hose ignited. Brookhaven National Laboratory, Lessons Learned 2002-CHBNL-MED-0003 (July 23, 2007).

 

BSC fire. Source: Stanford University,Use of open flames in Cabinets/Tissue Culture Hoods (May 29, 2003).


The following are examples of burners and heaters that could be used in a biosafety cabinet if other sterile techniques are not feasible:

  • Burner:  Touch-O-Matic Bunsen Burner
    Simply depress ON/OFF platform with side of hand to release gas stream, and continuous pinpoint pilot light ignites gas to produce full flame. Release platform and flame goes out to conserve gas. To produce continuous flame, depress platform, then turn it slightly; reverse process to turn off flame.
  • Heater:  Bacti-Cinerator
    Infrared heat chamber sterilizes loops, needles, and culture tubes in 5 to 7 seconds. Suitable for anaerobic procedures in chambers and hoods. Electric heat source eliminates hazards from gas and open flames. Within 6 minutes of activation, the interior of the ceramic cone reaches an optimum sterilizing temperature of 815°C (1,500°F). A prominent light indicates when the unit is in operation. Weighted cast aluminum stand includes handy spaces for storage of six inoculating loop handles. Electrical: 120V, 50/60Hz. UL listed. Unit is not intended for use with scalpels, forceps, or sharp instruments.
image   Text Box:
     

Touch-O-Matic Bunsen Burner.
Source: Fisher Scientific (May 2010)
.

 

Bacti-Cinerator
Source: VWR (May 2010)..


Aspirator Bottles or Suction Flasks
. Aspirator bottles or suction flasks should be connected to an overflow collection flask containing appropriate disinfectant, and to an in-line HEPA or equivalent filter (see Figure 9). This combination will provide protection to the central building vacuum system or vacuum pump, as well as to the personnel who service this equipment. Inactivation of aspirated materials can be accomplished by placing sufficient chemical decontamination solution into the flask to inactivate the microorganisms as they are collected. Once inactivation occurs, liquid materials can be disposed of as noninfectious waste.

Aspirator bottles that collect Risk Group (RG) 1 or RG2 biological materials that do not contain RG2 infectious agents may be placed outside the BSC as long as the aspirator bottles are placed inside a secondary spill tray.


Figure 9. Aspiration and House Vacuum System Protection. Shown below is one method to protect a house vacuum system during aspiration of infectious fluids. The left suction flask (A) is used to collect the contaminated fluids into a suitable decontamination solution; the right flask (B) serves as a fluid overflow collection vessel. An in-line HEPA filter (C) is used to protect the vacuum system (D) from aerosolized microorganisms. A spill tray (E) should be used when the flasks are outside the BSC. Source: adapted from BMBL, fifth edition, Appendix A.

E.4    Biosafety Cabinet Decontamination and Moves

E.4.1    Cabinet Surface Decontamination

Cabinet Surfaces. With the cabinet blower running, all containers and equipment should be surface decontaminated and removed from the cabinet when work is completed. At the end of the workday, the final surface decontamination of the cabinet should include a wipe-down of the work surface, the cabinet's sides and back, and the interior of the glass. If necessary, the cabinet should also be monitored for radioactivity and decontaminated when necessary. Investigators should remove their gloves and gowns in a manner to prevent contamination of unprotected skin and aerosol generation, and wash their hands as the final step in safe microbiological practices. The cabinet blower may be turned off after these operations are completed, or it may be left on.

Small Spills. Small spills within the operating BSC can be handled immediately by removing the contaminated absorbent paper towel and placing it into the biohazard bag or receptacle. Any splatter onto items within the cabinet, as well as the cabinet interior, should be immediately cleaned up with a towel dampened with an appropriate decontaminating solution. Gloves should be changed after the work surface is decontaminated and before placing clean absorbent towel in the cabinet. Hands should be washed whenever gloves are changed or removed.

Large Spills. Spills large enough to result in liquids flowing through the front or rear grilles require more extensive decontamination. All items within the cabinet should be surface decontaminated and removed. After ensuring that the drain valve is closed, decontaminating solution can be poured onto the work surface and through the grille(s) into the drain pan.

Decontamination Time and Cleanup. Twenty to 30 minutes is generally considered an appropriate contact time for decontamination, but this varies with the disinfectant and the microbiological agent. Manufacturer's directions should be followed. The spilled fluid and disinfectant solution on the work surface should be absorbed with paper towels and discarded into a biohazard bag. The drain pan should be emptied into a collection vessel containing disinfectant. A hose barb and flexible tube should be attached to the drain valve and be of sufficient length to allow the open end to be submerged in the disinfectant within the collection vessel. This procedure serves to minimize aerosol generation. The drain pan should be flushed with water and the drain tube removed.

Radioactive Materials. Should the spilled liquid contain radioactive material, a similar procedure can be followed. Radiation safety personnel should be contacted for specific instructions.

Work Surface, Grille, and Drain Pan Cleaning. Periodic removal of the cabinet work surface and/or grilles after the completion of drain pan decontamination may be justified because of dirty drain pan surfaces and grilles, which ultimately could occlude the drain valve or block airflow. However, extreme caution should be observed while wiping these surfaces to avoid injury from broken glass and sharp metal edges. Always use disposable paper towels and avoid applying harsh force. Wipe dirty surfaces gently. Never leave paper towels on the drain pan because the paper could block the drain valve or the air passages in the cabinet.

E.4.2 Internal Cabinet Gaseous Decontamination

BSCs that have been used for work involving infectious materials must be decontaminated before HEPA filters are changed or internal repair work is done. Before a BSC is relocated, a risk assessment considering the agents manipulated within the BSC must be performed to determine the need and method for decontamination. LBNL policy requires that BSCs and their filters be decontaminated with a gaseous decontaminant prior to being moved or internal repair work is conducted, unless approved by the Biosafety Officer. The most common decontamination method uses formaldehyde gas, although more recently, hydrogen peroxide vapor and chlorine dioxide gas have been used successfully.

E.5    Biosafety Cabinet Installation and Engineering

Room Ventilation and Secondary Barriers. Whereas BSCs are considered to be the primary safety barrier for manipulation of infectious materials, the laboratory room itself is considered to be the secondary safety barrier. Inward directional airflow is established by exhausting a greater volume of air than is supplied to a given laboratory and by drawing makeup air from the adjacent space. This directional air flow into the room should generally be accomplished at BL2 (see Section 5.6.4.1 of this manual). The air balance for the entire facility should be established and maintained to ensure that air flows from areas of least to greatest potential contamination.

The room exhaust system should be sized to handle both the room and all containment devices vented through the system. Adequate supply air must be provided to ensure appropriate function of the exhaust system. The facility engineer must be consulted before locating a new cabinet requiring connection to the building exhaust system. Right angle bends, long horizontal runs, and transitional connections within the systems will add to the demand on the exhaust fan. The building exhaust air should be discharged away from supply air intakes to prevent reentrainment of laboratory exhaust air into the building air supply system. Refer to recognized design guides for locating the exhaust terminus relative to nearby air intakes.

Utility Services. Utility services needed within a BSC must be planned carefully. Protection of vacuum systems must be addressed (Figure 9). Electrical outlets inside the cabinet must be protected by ground fault circuit interrupters and should be supplied by an independent circuit. When propane or natural gas is provided, a clearly marked emergency gas shutoff valve must be installed outside the cabinet for fire safety. All nonelectrical utility services should have exposed, accessible shutoff valves. The use of compressed air within a BSC must be carefully considered and controlled to prevent aerosol production and reduce the potential for vessel pressurization.

Ultraviolet (UV) Lamps. UV lamps are not required in BSCs nor are they necessary. If installed, UV lamps must be cleaned weekly to remove any dust and dirt that may block the germicidal effectiveness of the ultraviolet light. The lamps should be checked weekly with a UV meter to ensure that the appropriate intensity of UV light is being emitted. UV lamps must be turned off when the room is occupied to protect eyes and skin from UV exposure, which can burn the cornea and cause skin cancer. If the cabinet has a sliding sash, close the sash when operating the UV lamp.

BSC Placement. BSCs were developed as workstations to provide personnel, environmental, and product protection during the manipulation of infectious microorganisms. Certain considerations must be met to ensure maximum effectiveness of these primary barriers. Whenever possible, adequate clearance should be provided behind and on each side of the cabinet to allow easy access for maintenance and to ensure that the cabinet air recirculated to the laboratory is not hindered. A 12- to 14-inch clearance above the cabinet may be required to provide for accurate air velocity measurement across the exhaust filter surface and for exhaust filter changes. When the BSC is hard ducted or connected by a canopy unit to the ventilation system, adequate space must be provided so that the configuration of the duct work will not interfere with airflow. The canopy unit must provide adequate access to the exhaust HEPA filter for testing.

The ideal location for the biological safety cabinet is away from the entry (i.e., the rear of the laboratory away from traffic), since people walking parallel to the face of a BSC can disrupt the air curtain. The air curtain created at the front of the cabinet is quite fragile, amounting to a nominal inward and downward velocity of 1 mph. Open windows, air supply registers, portable fans, or laboratory equipment that creates air movement (e.g., centrifuges, vacuum pumps) should not be located near the BSC. Similarly, chemical fume hoods must not be located close to BSCs.

E.6    Biosafety Cabinet Testing and Certification

Class II BSCs are the primary containment devices that protect the worker, product, and environment from exposure to microbiological agents. BSCs used for BL1, BL2, or other safety levels must be tested and certified before initial use, after being moved, and on a nominal one-year cycle. This testing must verify that BSC operation is in accordance with the National Sanitation Foundation (NSF)/ American National Standard (ANSI)49 Standard (Class II Laminar Flow Biohazard Cabinetry) and be performed by experienced and qualified personnel. This testing ensures the balance of inflow and exhaust air, distribution of air onto the work surface, integrity of the cabinet and the filters, and other BSC features. The LBNL Environment, Health, and Safety (EH&S) Industrial Hygiene Group manages surveys and tests of BSCs through the LBNL ventilation safety program and qualified vendors contracted to test BSCs (see Section 5.6.4.2 of this manual).