Office of Health and Safety

Primary Containment for Biohazards

SECTION VII

The evolution of containment equipment for varied research and diagnostic applications created the need for consistency in construction, certification and performance. A Federal standard was developed⁰ to establish classes of air cleanliness and methods for monitoring clean work stations and clean rooms where HEPA filters are used to control airborne particulates.

The first "standard" to be developed specifically for BSCs¹⁷ served as a Federal procurement specification for the NIH Class II, Type 1 (now called Type A) biological safety cabinet, which had a fixed or hinged front window or a vertical sliding sash, vertical downward laminar airflow and HEPA-filtered air supply and exhaust. This guideline specified design criteria and defined prototype tests for microbiological aerosol challenge, velocity profiles, and leak testing of the HEPA filters. A similar procurement specification was generated²⁰ when the Class II Type 2 (now called Type B1) cabinet was developed.

The National Sanitation Foundation (NSF International) Standard No. 49 for Class II (Laminar Flow) Biohazard Cabinetry²⁴ was first published in 1976, providing the first independent standard for design, manufacture and testing the BSCs. This standard "replaced" the NIH specifications which were being used by other institutions and organizations purchasing BSCs. NSF Standard 49 incorporates specifications regarding design, materials and construction. This Standard for biological safety cabinets establishes performance criteria and provides the minimum requirements that are accepted in the United States. Cabinets which meet the standard and are certified by the NSF bear an NSF 49 Seal.

Standard No. 49 pertains to all models of Class II cabinets (Type A, B1, B2, and B3) and lists a series of specifications regarding:

• design/construction,

• performance,

• installation recommendations,

• recommended microbiological decontamination procedure, and

• references and specifications pertinent to Class II Biohazard Cabinetry.

While the NSF standard does not cover field testing of BSCs, it is common for many of its test methods and parameters to be applied in the field, and these are included in Annex "F" of the standard. Most recently revised in 1992 (with a new revision due in 2000),²⁴ this Standard is reviewed periodically by a steering committee to ensure that it remains consistent with developing technologies.⁰

The operational integrity of a new BSC must be validated before it is put into service or after a cabinet has been repaired or relocated. Relocating a BSC may break the HEPA filter seals or otherwise damage the filters or the cabinet. Each BSC should be tested and certified at least annually to ensure continued proper operation.

On-site testing following the recommendations for field testing (NSF Standard 49) must be performed by experienced, qualified personnel. Some basic information is included here to assist in understanding the frequency and kinds of tests to be performed. In 1993, NSF began a program for accreditation of certifiers based on written and practical examinations. Education and training programs for persons seeking accreditation as qualified to perform all field certification tests are offered through the Eagleson Institute, Sanford, ME; the Harvard School of Public Health, Cambridge, MA; NuAire Inc., Plymouth, MN; Forma Scientific Inc., Marietta, OH; and Lab Conco Corporation, Kansas City, MO. Other training, education and certification programs may be developed in the future. Selecting competent individuals to perform testing and certification is important, and it is suggested that the institutional biosafety officer be consulted in identifying companies qualified to conduct the necessary field performance tests.

It is strongly recommended that whenever possible accredited field certifiers be used to test and certify BSCs If in-house personnel are preforming the certifications, then these individuals should become accredited. The importance of proper certification cannot be emphasized enough, since persons who manipulate infectious microorganisms are at increased risk of acquiring an occupational illness when their BSCs are functioning improperly.

The annual tests applicable to each of the three classes of biological safety cabinets are listed in Table 3. Table 4 indicates where to find information regarding the conduct of selected tests. BSCs perform consistently well when proper annual certification procedures are followed; cabinet or filter failures tend to occur infrequently.

BSCs are the primary containment device that protect the worker, product and environment from exposure to microbiological agents. Their operation as specified by Standard No. 49 needs to be verified at the time of installation and annually thereafter. The purpose and acceptance level of the performance tests (Table 3) are to ensure the balance of inflow and exhaust air, the distribution of air onto the work surface, and the integrity of the cabinet. Other tests check electrical and physical features of the BSC.

This test is performed to measure the velocity of air moving through the cabinet workspace, and is to be performed on all biosafety cabinets.

This test is performed to determine the calculated or directly measured velocity through the work access opening, to verify the nominal set point average inflow velocity and to calculate the exhaust airflow volume rate.

This test is performed to determine if the airflow along the entire perimeter of the work access opening is inward, if airflow within the work area is downward with no dead spots or refluxing, if ambient air passes onto or over the work surface, and if there is refluxing to the outside at the window wiper gasket and side seals. The smoke test is an indicator of airflow direction, not velocity.

This test is performed to determine the integrity of supply and exhaust HEPA filters, filter housing, and filter mounting frames while the cabinet is operated at the nominal set point velocities. An aerosol in the form of generated particulates of dioctylphthalate (DOP) or an accepted alternative (e.g., food grade corn oil, di(2-ethylhexyl), sebecate, polyethylene glycol, and medical grade light mineral oil) is required for leak-testing HEPA filters and their seals. Although DOP has been identified as a potential carcinogen, competent service personnel are trained to use this chemical in a safe manner. The aerosol is generated on the intake side of the filter, and particles passing through the filter or around the seal are measured with a photometer on the discharge side. This test is suitable for ascertaining the integrity of all HEPA filters.

The pressure holding test is performed to determine if exterior surfaces of all plenums, welds, gaskets, and plenum penetrations or seals are free of leaks. It need only be performed just prior to initial installation when the BSC is in a free-standing position (all four sides are easily accessible) in the room in which it will be used, after a cabinet has been relocated to a new location, and again after removal of access panels to plenums for repairs or a filter change. This test may also be performed on fully installed cabinets. "Cabinet integrity can also be checked using the bubble test "

These safety tests are performed to determine if a potential shock hazard exists by measuring the electrical leakage, polarity, ground fault interrupter function, and ground circuit resistance to the cabinet connection. They may be performed by an electrical technician other than the field certification personnel at the same time the other field certification tests are conducted. The polarity of electrical outlets are checked (see Table 3, E). The ground fault circuit interrupter should trip when approximately 5 milliamperes (ma) is applied.

This test is performed to measure the light intensity on the work surface of the cabinet as an aid in minimizing cabinet operator’s fatigue.

This test is performed to determine the amount of vibration in an operating cabinet as a guide to satisfactory mechanical performance, as an aid in minimizing cabinet operator's fatigue, and to prevent damage to delicate tissue culture specimens.

This test is performed to measure the noise levels produced by the cabinets, as a guide to satisfactory mechanical performance and an aid in minimizing cabinet operator's fatigue.

: A few BSCs have UV lamps. When used, they must be tested periodically to ensure that their energy output is sufficient to kill microorganisms. After having been turned off and allowed to cool, the surface on the bulb should be cleaned with 70% ethanol prior to performing this test. Five minutes after the lamp has been turned on, the sensor of the UV meter is placed in the center of the work surface. The radiation output should not be less than 40 microwatts per square centimeter at 254 nanometers (nm).

Finally, accurate test results can only be assured when the testing equipment is properly maintained and calibrated. It is appropriate to request the calibration information for the test equipment being used by the certifier.

This page last reviewed May 17, 2001

Office of the Director/Administrator
Centers for Disease Control and Prevention
and Agency for Toxic Substances and Disease Registry
Atlanta, GA