Influence of crossdrafts on the performance of a biological safety cabinet.

A biological safety cabinet was tested to determine the effect of crossdrafts (such as those created by normal laboratory activity or ventilation) upon the ability of the cabinet to protect both experiments and investigators. A simple crossdraft, controllable from 50 to 200 feet per min (fpm; 15.24 to 60.96 m/min), was created across the face of the unit. Modifications of standardized procedures involving controlled bacterial aerosol challenges provided stringent test conditions. Results indicated that, as the crossflow velocities exceeded 100 fpm, the ability of the cabinet to protect either experiments or investigators decreased logarithmically with increasing crossdraft speed. Because 100 fpm is an airspeed easily achieved by some air conditioning and heating vents (open windows and doorways may create velocities far in excess of 200 fpm), the proper placement of a biological safety cabinet within the laboratory--away from such disruptive air currents--is essential to satisfactory cabinet performance.     

Microbiological evaluation of a biological safety cabinet modified for bedding disposal

A biological safety cabinet modified for bedding disposal was tested to determine the cabinet's ability to protect operators and experiments from aerosol exposure during routine microbiological and cage cleaning procedures. Stringent test conditions were provided by modifications of standardized protocols in addition to simulated cage dumping procedures, both of which utilized bacterial aerosols as challenges. Results of standardized test procedures (with no operator present) indicated good performance in protecting both operators and experiments. Procedures involving the dumping (by an operator) of contaminated bedding within the unit showed that the cabinet was able to contain 99.96% or greater of the total particles generated.     

Containment of Microbial Aerosols in a Microbiological Safety Cabinet

A microbiological safety cabinet was evaluated to determine conditions under which microorganisms might escape. Tests were conducted under three cabinet-closure conditions, various airflow velocities, and different laboratory operations, with 10(5), 1.1 x 10(5), and 10(6) microorganisms per cubic foot of cabinet space released per min for 5 min. The data revealed that (i) escape of a human infectious dose is possible when the cabinet is used with the glove panel off; (ii) the number of organisms that escaped from the cabinet increased with a decrease in air velocity; and (iii) an increase in the number of laboratory operations resulted in an increase in the number of organisms that escaped. Thus, when the glove panel was off, the cabinet was only safe for operations that released a small number of microorganisms into the cabinet, whereas the cabinet was safe for operations of significantly greater hazard when used with the glove panel on but with the gloves unattached.     

国内生物安全柜产品标准（JG170和YY0569）比较

国内生物安全柜执行的两份标准（ JG170-2005和YY0569-2011）存在一定差异,导致实际执行中存在分歧和疑惑。该文就两份标准中对生物安全柜的结构要求,性能要求和测试方法进行了比较,并对其逐一进行客观分析解释,对生物安全柜在实际维护和检验时提供了指导。     

Modified Laminar Flow Biological Safety Cabinet

Tests are reported on a modified laminar flow biological safety cabinet in which the return air plenum that conducts air from the work area to the high efficiency particulate air filters is under negative pressure. Freon gas released inside the cabinet could not be detected outside by a freon gas detection method capable of detecting 10(-6) cc/s. When T3 bacteriophage was aerosolized 5 cm outside the front opening in 11 tests, no phage could be detected inside the cabinet with the motor-filter unit in operation. An average of 2.8 x 10(5) plaque-forming units (PFU)/ft(3) (ca. 0.028 m(3)) were detected with the motor-filter unit not in operation, a penetration of 0.0%. Aerosolization 5 cm inside the cabinet yielded an average of 10 PFU/ft(3) outside the cabinet with the motor-filter unit in operation and an average of 4.1 x 10(5) PFU/ft(3) with the motor-filter unit not in operation, a penetration of 0.002%. These values are the same order of effectiveness as the positive-pressure laminar flow biological safety cabinets previously tested. The advantages of the negative-pressure return plenum design include: (i) assurance that if cracks or leaks develop in the plenum it will not lead to discharge of contaminated air into the laboratory; and (ii) the price is lower due to reduced manufacturing costs.     

Microbiological Studies on the Performance of a Laminar Airflow Biological Cabinet

Engineering and microbiological tests indicated that a typical, commercial laminar airflow cabinet was not effective in providing either product protection or agent containment. The cabinet was modified and tested through a series of alternate configurations to establish a set of design criteria. A mock-up cabinet was developed from these design criteria. The mock-up unit was evaluated for efficiency in providing both product protection and agent containment. In these evaluations, challenge methods were developed to simulate normal, in-use laboratory operations. Controlled bacterial or viral aerosol challenges were used at higher than normal levels to provide stringent test conditions. Test results indicated that the mock-up unit was considerably better in preventing agent penetration (0.1 to 0.2 particles per 100 ft(3) of air) than the commercial cabinet (5 to 6 particles per 100 ft(3) of air) during product protection tests. Similarly, agent containment was considerably better in the new cabinet (particle escape of 2 to 3 per 100 ft(3) of air at only one of the five test sites) than in the commercial cabinet (particle escape of 2 to 14 per 100 ft(3) of air at three of the five test sites).     

Bacteriological testing of a modified laminar flow microbiological safety cabinet

A modified microbiological safety cabinet which can be used as a class II and a class III safety cabinet has been bacteriologically tested. This cabinet makes use of a high-speed down-flow air curtain in the front opening to minimize the amount of air escaping over the arms of the operator. By using artificial aerosols and a dummy or a test person placing his arms into the working opening of the cabinet, a transfer from the inside to the environment was detected only when the highest concentration of the test aerosol was used. Since the number of bacteria detected was very low, this is considered to be acceptable. when the cabinet was used as a class III type, with a glove panel mounted in the front opening, leakage from the environment occurred. This could be completely prevented by fixing tape over the hinge of the front panel.The conclusion is drawn that this type of biohazard hood can be safely used as a class II and a class III microbiological safety cabinet, provided the construction of the hinge of the front panel will be adapted to prevent transfer from the environment to the working area.     

Biological Containment Facility for Studying Infectious Disease

To effectively characterize newly recognized viruses (Marburg, Lassa, etc.) and to study other highly virulent infections for which no effective prophylaxis or therapy exists, special containment facilities must be utilized and conventional techniques modified to minimize risk to laboratory personnel. This paper describes a laboratory suite for such studies, contained within a larger research facility; two separate biological safety cabinet systems, animal rooms support laboratories, change room facilities, shower, air lock, and other safety features are contained in the area. Details of design, construction, airflows, and equipment are described in addition to a discussion of operation, techniques, and modification of laboratory equipment utilized in actual studies.     

Improving the biosafety of cell sorting by adaptation of a cell sorting system to a biosafety cabinet.

BACKGROUND: The jet-in-air cell sorters currently available are not very suitable for sorting potentially biohazardous material under optimal conditions because they do not protect operators and samples as recommended in the guidelines for safe biotechnology. To solve this problem we have adapted a cell sorting system to a special biosafety cabinet that satisfies the requirements for class II cabinets. With aid of this unit, sorting can be performed in conformance with the recommendations for biosafety level 2. METHODS: After integrating a modified fluorescence-activated cell sorter (FACS) Vantage into a special biosafety cabinet, we investigated the influence of the laminar air flow (LAF) inside the cabinet on side stream stability and the analytical precision of the cell sorter. In addition to the routine electronic counting of microparticles, we carried out tests on the containment of aerosols, using T4 bacteriophage as indicators, to demonstrate the efficiency of the biosafety cabinet for sorting experiments performed under biosafety level 2 conditions. RESULTS: The experiments showed that LAF, which is necessary to build up sterile conditions in a biosafety cabinet, does not influence the conditions for side stream stability or the analytical precision of the FACS Vantage cell sorting system. In addition, tests performed to assess aerosol containment during operation of the special biosafety cabinet demonstrated that the cabinet-integrated FACS Vantage unit (CIF) satisfies the conditions for class II cabinets. In the context of gene transfer experiments, the CIF facility was used to sort hematopoietic progenitor cells under biosafety level 2 conditions. CONCLUSIONS: The newly designed biosafety cabinet offers a practical modality for improving biosafety for operators and samples during cell sorting procedures. It can thus also be used for sorting experiments with genetically modified organisms in conformance with current biosafety regulations and guidelines.     

Efficient Steady-State Volatile Organic Compound Removal from Air by Live Bacteria Immobilized on Fiber Supports

Fibers are suggested for bacterial immobilization in trickle-bed bioreactors used for the removal of volatile organic compounds (VOCs) from air. Fiber-based bioreactors retain up to 200 to 300 mg of dry biomass per 1 g of support, which is a much larger value than that of traditional, granule-based bioreactors. Air pollutant removal efficiency for fiber-based bioreactors remains high with large inlet pollutant concentrations or space velocities (lower contact times). Efficient removal is achieved not only for a water-miscible substrate (ethanol), but also for some less water-soluble compounds, such as ethyl acetate and styrene. Specific pollutant elimination capacity per unit fiber-based biocatalyst volume (up to 4000 g/m³-h) exceeds those of biological air purification methods and is comparable to chemical methods. Unlike granule-based biocatalysts, oxygen limitation for pollutant biodegradation is not observed. Evidence obtained shows that the higher air purification efficiency is due to the greater surface-to-volume ratio of fibers when compared with granules, which results in a more efficient substrate mass transfer.     

Resolution as a function of accelerating voltage in electron microscopy of semithick biological specimens

In the past, biological sections ranging in thickness from 0.10- to 0.50-micron have usually been examined with high-voltage (greater than 500 kV) electron microscopes (HVEM). Now investigators are increasingly using intermediate voltage (200-500 kV) electron microscopes (IVEM), which are more readily available and demand less maintenance. In a study of "typical" plastic-embedded, stained sections of mouse liver ranging from 0.10 to 1.0 micron thick, we determined the resolution obtainable at 100, 200, and 1000 kV. At all three accelerating voltages the resolution (2.7 nm) for 0.10-micron sections was limited only by the sections stain granularity. For 0.25-micron thickness the resolutions were 5.8, 3.1, and 3.1 nm at 100, 200, and 1000 kV, respectively. The maximum usable thickness at 200 kV with resolution sufficient to resolve membranes clearly was between 0.75 and 1.0 micron, depending on the magnification. Resolution at 100 kV was adequate for screening sections up to 1.0-micron thick for preparation defects prior to examination with an IVEM or HVEM.     

A Safe Method of Exposing Microthreads in the Open Air

A safe method for exposing pathogenic organisms held on microthreads to realistic environmental conditions is described. The apparatus consists of an air duct in which the organisms on microthreads are exposed, and contiguous to it a safety cabinet, air from both systems being continuously extracted into high efficiency aerosol filters.     

New donors and acceptors of nitrogen oxide as a potential therapeutical agents

The synthesis of new donors and acceptors of nitrogen oxide is described. New lipophilic nitronylnitroxyl radicals (NNR) that act as paramagnetic scavengers of nitrogen oxide are synthesized and characterized. The purity of the preparations is determined, and their structures are confirmed. The lipophilicity of the radicals is tested by ESR spectroscopy. The incorporation into lipid multilayers is shown to protect NNR from reduction in biological samples, while their ability to scavenge nitrogen oxide and form iminonitroxyl radicals is retained. A decreased rate of NNR reduction under these conditions substantially enhances their effectiveness as paramagnetic acceptors of nitrogen oxide in biological systems. The synthesis of a new hydrophilic NO donor, 3-bromo-3,4-dihydro-4,4-dimethyl-4-(2-pyridyl)-diazet-1,2-dioxide (DDpyr), is described. The constants of DDpyr decomposition in tris-HCl buffer (pH 7.5) and in DMSO are determined (4.5 x 10(-6) and 0.5 x 10(-6) s-1, respectively). A substantially higher rate of of DDpyr decomposition in buffer, compared with the decomposition rates determined previously for some diazetines, makes DDpyr a prospective candidate for the use in aqueous media. It is found in experiments on perfused rat caudal artery that DDpyr is an effective vasodilator. Intraperitoneal injection of DDpyr to hereditarily hypertensive rats (ISIAH line) at doses of 100-200 micrograms/kg body mass considerably diminishes their systolic arterial pressure.     

Irreversible thermoinactivation of ribonuclease‐A by soft‐hydrothermal processing

Ribonuclease (RNase), which often represents molecular biological contamination, is a thermostable enzyme. When RNase is heated at 121°C by autoclave sterilization for 20 min, it does not lose its activity. However, the nature of the molecular events by which the irreversible denaturation occurs remains unknown. The purpose of this study was to elucidate the molecular mechanisms of irreversible thermal denaturation of RNase A and to develop an advanced sterilization method using soft‐hydrothermal processing, which has the advantages of improved safety and cost‐efficiency. The enzymatic activity of RNase was measured using polyacrylamide gel electrophoresis with torula yeast RNA. We evaluated the temperature and time course of irreversible thermoinactivation of RNase by normal autoclaving, hot‐air sterilization, and soft‐hydrothermal processing that had been controlled to the desired steam saturation ratio. The results indicated that RNase A was deactivated by autoclave sterilization (121°C, 20 min) immediately after treatment, but was reactivated over time. Hot‐air sterilization (180°C, atmospheric pressure, 60 min) produced results similar to that of autoclave sterilization. In contrast, RNase A was irreversibly thermoinactivated by soft‐hydrothermal processing (110°C, 20 min) at 100% steam saturation ratio. We also determined that the mechanism of irreversible thermoinactivation of RNase A involved hydrolysis and deamidation under this condition at a steam saturation ratio of more than 100%. © 2009 American Institute of Chemical Engineers Biotechnol. Prog., 2009     

Profiling gene expression using onto-express

Gene expression profiles obtained through microarray or data mining analyses often exist as vast data strings. To interpret the biology of these genetic profiles, investigators must analyze this data in the context of other information such as the biological, biochemical, or molecular function of the translated proteins. This is particularly challenging for a human analyst because large quantities of less than relevant data often bury such information. To address this need we implemented an automated routine, called Onto-Express (http://vortex.cs.wayne.edu:8080), to systematically translate genetic fingerprints into functional profiles. Using strings of accession or cluster identification numbers, Onto-Express searches the public databases and returns tables that correlate expression profiles with the cytogenetic locations, biochemical and molecular functions, biological processes, cellular components, and cellular roles of the translated proteins. The profiles created by Onto-Express fundamentally increase the value of gene expression analyses by facilitating the translation of quantitative value sets to records that contain biological implications.     

Mass Airflow Cabinet for Control of Airborne Infection of Laboratory Rodents

A mass airflow cabinet for handling and housing of laboratory rodents has been developed and tested. The unit consists of a high-efficiency particulate air filter and uniform distribution of air at a vertical velocity of 19 cm per s. Animals are maintained without bedding in mesh-bottomed cages that rest on rollers for rotation inside the cabinet. There is an air barrier of 90 cm per s separating the cabinet air from room air. Sampling for airborne bacteria yielded an average of 0.03 colony-forming units (CFU) per ft(3) of air inside the cabinet, whereas 28.8 CFU per ft(3) was simultaneously detected outside the cabinet during housekeeping, a reduction of almost three logs. The efficiency of the air barrier was tested by aerosolization of T3 phage. When phage was aerosolized 5 cm outside the cabinet, no phage could be detected 5 cm inside when the fans were operating; with the fans off an average of 1.6 x 10(4) plaque-forming units (PFU) per ft(3) was detected in six tests. Aerosolization of phage inside the cabinet yielded an average of 9 x 10 PFU per ft(3) outside; an average of 4.1 x 10(6) PFU per ft(3) were detected with the fans not in operation, a reduction of more than four logs. In-use studies on effectiveness showed that the cabinet significantly reduced the incidence of mice originally titer-free to Reo-3 virus. Hemagglutination inhibition antibodies to Reo-3 were detected in 9/22 (42%) mice housed in a conventionally ventilated animal laboratory while no seroconversion was detected in any of 22 mice housed in the mass air flow cabinet in the same laboratory.     

Formaldehyde degradation by catalytic oxidation.

Formaldehyde used for the disinfection of a laminar-flow biological safety cabinet was oxidatively degraded by using a catalyst. This technique reduced the formaldehyde concentration in the cabinet from about 5,000 to about 45 mg/m3 in 8 h. This technique should prove useful in other applications.     

A hollow-fiber membrane bioreactor for the removal of trichloroethylene from the vapor phase

A hollow-fiber membrane bioreactor was used to separate trichloroethylene (TCE) from a gaseous waste stream with subsequent cometabolic biodegradation by a pure culture of Methylosinus trichosporium OB3b PP358. The two-stage bioreactor system was successfully operated for 20 days. PP358 was grown in a continuous-flow chemostat and circulated through the fiber lumen of a hollow-fiber membrane module (HFMM), while TCE contaminated air (141 to 191 microg/L) was pumped through the HFMM shell. Between 54% -84% TCE transfer and 92%-96% TCE cometabolism were obtained in the HFMM reactor loop. Short shell-residence times, 1.6 to 5.0 minutes, demonstrated quick throughput of TCE contaminated air. Best-fit computer modeling of the biological experiments estimated mass transfer coefficients between 2.0 x 10(-3) cm/min and 5.6 x 10(-3) cm/min. The average pseudo-first-order biodegradation rate constant for the biological experiments was 0.46 L/mg TSS/d. These results demonstrate that the hollow-fiber membrane bioreactor represents an attractive technology for the bioremediation of gaseous waste streams.     

Open fronted safety cabinets in ventilated laboratories

Open fronted Class I and II microbiological safety cabinets (MSCs) are required by the British Standard 5726 to provide similar levels of operator protection (viz. 10⁵). In laboratories that are naturally ventilated large numbers of both types of cabinets have been shown to exceed this requirement consistently over a number of years. The designs of some mechanically ventilated laboratories, however, produce excessive turbulence and draughts that can prejudice containment at the front aperture. On-site commissioning tests to determine operator protection factor are now well established and are recognized as being essential to the setting up of all open fronted cabinets in both ventilated and unventilated laboratories. This paper shows that where environmental conditions induce unsatisfactory cabinet containment, adjustments to air supply and exhaust systems can be made which will enable both Class I and II cabinets to produce operator protection factors in excess of 10⁵. When compatibility is achieved between the local environment and the cabinets it is demonstrated that disturbances at the front aperture, caused by operator working procedures or by disturbances due to personnel movement within the room, have similar effects on both Class I and II cabinets. Once performance levels have been satisfactorily achieved, regular containment testing has shown that consistent performance can be maintained. These aspects of open fronted safety cabinet performance are discussed in relation to ventilated laboratories suitable for work with the human immunodeficiency virus (HIV). Of paramount importance in the future is the necessity to design laboratory air systems that will be compatible with satisfactory safety cabinet performance—a relatively new requirement in ventilation system specifications.     

Effects of different stressor agents on gilthead seabream natural cytotoxic activity

Several common situations in gilthead seabream (Sparus aurata L.) farming, such as air exposure, crowding and the use of anaesthetics, have been demonstrated to be stressful. In the present study, these conditions were simulated in the laboratory, after which head-kidney natural cytotoxic cell (NCC) activity was evaluated. For this, several specimens were air exposed for 2 min, returned to the aquarium and sampled from 0 to 4 days after exposure. NCC activity was significantly lower on the day following air-exposure compared with the control (rested fish) but not at any other time studied. Other fish were crowded (100 kg biomass m(-3), 2 h), returned to an aquarium with the same density as the control group (9 kg m(-3)) and sampled from 0 to 4 days after treatment. Head-kidney NCC activity was statistically increased compared with the control (resting) fish, 1 day after crowding. Anaesthesis for 1 h with 60 or 200 microl 2-phenoxyethanol l(-1)had no significant effect on NCC activity, while the use of 50 mg MS222 l(-1)for 1 h reduced such activity (by about 40%) compared with the control. In other experiments, fish were consecutively treated with crowding and anaesthetics. When treated with the lowest 2-phenoxyethanol concentration after crowding, the NCC activity inhibition was abolished compared with the activity in fish treated either with crowding or anaesthetic alone, while the use of the highest concentration increased such inhibition. The use of MS222 after crowding did not produce any differential effect compared with the fish treated with only one of the factors. In conclusion, NCC activity is affected differently according to the stress factor applied (hypoxia, crowding and/or anaesthetics). Differences in the effects provoked by these stressors on other seabream innate immune parameters are discussed.