A biosafety cabinet (BSC) is an enclosed, ventilated laboratory workstation designed to contain biological hazards, providing a barrier between the worker and the materials being handled. The primary function of a BSC is to ensure protection for three elements: the laboratory personnel, the product or sample being manipulated, and the surrounding environment. This triple protection is achieved through carefully controlled, high-efficiency filtered airflow patterns. The standardization of these devices by organizations like the Centers for Disease Control and Prevention (CDC) has made the BSC an indispensable tool for safe biological research.
Classification of Biosafety Cabinets
Biosafety cabinets are categorized into three main classes—Class I, Class II, and Class III—based on their construction, airflow mechanism, and the degree of protection they provide. The selection of the appropriate class depends directly on the biological risk level of the agents being used. All three classes provide protection for the personnel and the environment, but only Class II and Class III also protect the product inside the cabinet from external contamination.
A Class I cabinet is the simplest type, drawing laboratory air inward through the front opening, across the work surface, and then filtering it through a High-Efficiency Particulate Air (HEPA) filter before exhausting it. This design provides robust protection for the user and the environment but offers no protection for the work or sample, as unfiltered room air flows directly over the work area. These cabinets are often used to enclose specific aerosol-generating equipment, such as centrifuges or vortex mixers, when product sterility is not a concern.
Class II cabinets are the most common type found in clinical and research laboratories because they offer simultaneous protection for the personnel, the environment, and the product. This triple protection is achieved by using both an inward airflow at the front opening and a vertical, downward-flowing curtain of HEPA-filtered air over the work surface. Class II cabinets are further divided into types A1, A2, B1, and B2, which differ primarily in the percentage of air that is recirculated versus exhausted, and the method of exhaust.
Type A1 and A2 cabinets recirculate a significant portion of the HEPA-filtered air back into the cabinet workspace, with the remainder exhausted back into the laboratory, also after passing through a HEPA filter. The A2 subtype is the most widely used due to its versatility and ability to handle Biosafety Level 1 through 3 agents. Because the exhaust air is returned to the room, A-type cabinets are not suitable for work involving volatile toxic chemicals or radionuclides, which the HEPA filter does not remove.
In contrast, Type B1 and B2 cabinets are designed for work involving biological agents in conjunction with volatile hazardous chemicals. Both B-types must be “hard-ducted,” meaning they are permanently connected to the building’s exhaust system, which vents the air outside. The Type B2 cabinet is a total exhaust system, meaning 100% of the air is pulled out and exhausted after HEPA filtration, with no air recirculated back into the work area. This complete exhaust mechanism makes the Type B2 the preferred choice for procedures generating significant amounts of volatile chemicals or radionuclides, as it ensures all chemical vapors are removed from the work zone.
The Class III cabinet represents the highest level of containment and is designed for work with the most dangerous biological agents, typically those requiring Biosafety Level 4 precautions. This cabinet is a completely enclosed, gas-tight system, often referred to as a glove box, where operations are conducted through attached, heavy-duty gloves. All materials entering or leaving the Class III cabinet must pass through a decontaminating chamber, such as a dunk tank or double-door autoclave, ensuring an airtight separation between the hazardous agent and the operator.
Principles of Airflow and Filtration
The protective function of a biosafety cabinet relies fundamentally on the precise control of airflow dynamics and the use of High-Efficiency Particulate Air (HEPA) filters. These filters are composed of a mat of randomly arranged borosilicate microfibers formed into pleated sheets to maximize the surface area for filtration. HEPA filters physically capture airborne particulates and microorganisms rather than chemically neutralizing them.
The effectiveness of a HEPA filter is rated by its ability to trap particles at the Most Penetrating Particle Size (MPPS), approximately 0.3 microns. At this size, the filter is certified to remove at least 99.97% of particles. Filtration occurs through three mechanical processes: interception, where a particle adheres to a fiber; impaction, where larger particles collide directly with the fibers; and diffusion, where tiny particles’ random movement increases their likelihood of contact and capture.
Airflow in a Class II BSC is engineered to create two distinct protective zones: the inflow and the downflow. The inward airflow, or inflow, is drawn through the front access opening at a specific velocity, creating an air barrier that prevents aerosols from escaping toward the user. This air is sucked through the front grille, under the work surface, and moves toward the blower, maintaining the cabinet under negative pressure relative to the room.
The downflow is a curtain of HEPA-filtered, sterile air that moves vertically across the work surface. This unidirectional, non-turbulent air stream protects the product from contamination by continuously flushing the work zone with clean air. The balance between the inflow velocity and the downflow velocity is calibrated to ensure the air barrier remains intact and containment is maintained.
Operational Safety and Decontamination
Safe and effective use of a biosafety cabinet depends on strict adherence to specific operational protocols that maintain the integrity of its airflow.
Operational Protocols
- Ensure the front sash is set to the manufacturer’s recommended working height (typically 8 to 10 inches) to preserve the protective air curtain.
- Minimize arm movements in and out of the cabinet, avoiding rapid motions that could disrupt the delicate airflow barrier.
- Never block the front and rear air intake grilles with equipment, supplies, or arms, as this compromises containment.
- Perform work at least four to six inches inside the front grille to allow the inflow air to fully capture aerosols.
- Allow the cabinet to run for several minutes before and after work to purge the air inside the work area.
To verify the cabinet’s performance, Biosafety Cabinets must undergo regular testing and certification by qualified technicians. This process is typically required annually or semi-annually and involves specialized checks, such as airflow velocity profiling and HEPA filter integrity testing, often following standards like NSF/ANSI 49. Certification is also mandatory after the cabinet has been moved or after a HEPA filter has been replaced to ensure the engineered containment features remain functional.
Decontamination procedures are divided into routine cleaning and full sterilization, depending on the need. Routine surface cleaning, performed before and after each use, involves wiping down the interior work surfaces with an approved liquid disinfectant, such as 70% ethanol. Full decontamination is a deeper process required before a cabinet is relocated, serviced internally, or discarded. This process typically involves gaseous fumigation using agents like formaldehyde or vaporized hydrogen peroxide (VHP), which sterilizes all internal surfaces and plenums, including the inaccessible areas around the filters.