Wayne Einfeld

Rapid Microchip-Based Electrophoretic Immunoassays For The Detection Of Swine Influenza Virus.

Towards developing rapid and portable diagnostics for detecting zoonotic diseases, we have developed microchip-based electrophoretic immunoassays for sensitive and rapid detection of viruses. Two types of microchip-based electrophoretic immunoassays were developed. The initial assay used open channel electrophoresis and laser-induced fluorescence detection with a labeled antibody to detect influenza virus. However, this assay did not have adequate sensitivity to detect viruses at relevant concentrations for diagnostic applications. Hence, a novel assay was developed that allows simultaneous concentration and detection of viruses using a microfluidic chip with an integrated nanoporous membrane. The size-exclusion properties of the in situ polymerized polyacrylamide membrane are exploited to simultaneously concentrate viral particles and separate the virus/fluorescent antibody complex from the unbound antibody. The assay is performed in two simple steps--addition of fluorescently labeled antibodies to the sample, followed by concentration of antibody-virus complexes on a porous membrane. Excess antibodies are removed by electrophoresis through the membrane and the complex is then detected downstream of the membrane. This new assay detected inactivated swine influenza virus at a concentration four times lower than that of the open-channel electrophoresis assay. The total assay time, including device regeneration, is six minutes and requires <50 microl of sample. The filtration effect of the polymer membrane eliminates the need for washing, commonly required with surface-based immunoassays, increasing the speed of the assay. This assay is intended to form the core of a portable device for the diagnosis of high-consequence animal pathogens such as foot-and-mouth disease. The electrophoretic immunoassay format is rapid and simple while providing the necessary sensitivity for diagnosis of the illness state. This would allow the development of a portable, cost-effective, on-site diagnostic system for rapid screening of large populations of livestock, including sheep, pigs, cattle, and potentially birds.

False-Negative Rate and Recovery Efficiency Performance of a Validated Sponge Wipe Sampling Method

ABSTRACT Recovery of spores from environmental surfaces varies due to sampling and analysis methods, spore size and characteristics, surface materials, and environmental conditions. Tests were performed to evaluate a new, validated sponge wipe method using Bacillus atrophaeus spores. Testing evaluated the effects of spore concentration and surface material on recovery efficiency (RE), false-negative rate (FNR), limit of detection (LOD), and their uncertainties. Ceramic tile and stainless steel had the highest mean RE values (48.9 and 48.1%, respectively). Faux leather, vinyl tile, and painted wood had mean RE values of 30.3, 25.6, and 25.5, respectively, while plastic had the lowest mean RE (9.8%). Results show roughly linear dependences of RE and FNR on surface roughness, with smoother surfaces resulting in higher mean REs and lower FNRs. REs were not influenced by the low spore concentrations tested (3.10 × 10−3 to 1.86 CFU/cm2). Stainless steel had the lowest mean FNR (0.123), and plastic had the highest mean FNR (0.479). The LOD90 (≥1 CFU detected 90% of the time) varied with surface material, from 0.015 CFU/cm2 on stainless steel up to 0.039 on plastic. It may be possible to improve sampling results by considering surface roughness in selecting sampling locations and interpreting spore recovery data. Further, FNR values (calculated as a function of concentration and surface material) can be used presampling to calculate the numbers of samples for statistical sampling plans with desired performance and postsampling to calculate the confidence in characterization and clearance decisions.

Diffusional sampler performance under transient exposure conditions

A laboratory exposure study was conducted with two commercially available diffusional samplers. Trichloroethylene exposures were carried out under both steady-state and transient concentration profiles to determine whether sampler bias exists under transient exposure conditions. Results indicate acceptable performance of diffusional samplers under both steady-state and transient exposure conditions.

Evaluation of surface sampling method performance for Bacillus Spores on clean and dirty outdoor surfaces.

Recovery of Bacillus atrophaeous spores from grime-treated and clean surfaces was measured in a controlled chamber study to assess sampling method performance. Outdoor surfaces investigated by wipe and vacuum sampling methods included stainless steel, glass, marble and concrete. Bacillus atrophaeous spores were used as a surrogate for Bacillus anthracis spores in this study designed to assess whether grime-coated surfaces significantly affected surface sampling method performance when compared to clean surfaces. A series of chamber tests were carried out in which known amounts of spores were allowed to gravitationally settle onto both clean and dirty surfaces. Reference coupons were co-located with test coupons in all chamber experiments to provide a quantitative measure of initial surface concentrations of spores on all surfaces, thereby allowing sampling recovery calculations. Results from these tests, carried out under both low and high humidity conditions, show that spore recovery from grime-coated surfaces is the same as or better than spore recovery from clean surfaces. Statistically significant differences between method performance for grime-coated and clean surfaces were observed in only about half of the chamber tests conducted.

False negative rate and other performance measures of a sponge-wipe surface sampling method for low contaminant concentrations.

Recovery of spores from environmental surfaces is known to vary due to sampling methodology, techniques, spore size and characteristics, surface materials, and environmental conditions. A series of tests were performed to evaluate a new, validated sponge-wipe method. Specific factors evaluated were the effects of contaminant concentrations and surface materials on recovery efficiency (RE), false negative rate (FNR), limit of detection (LOD) - and the uncertainties of these quantities. Ceramic tile and stainless steel had the highest mean RE values (48.9 and 48.1%, respectively). Faux leather, vinyl tile, and painted wood had mean RE values of 30.3, 25.6, and 25.5, respectively, while plastic had the lowest mean RE (9.8%). Results show a roughly linear dependence of surface roughness on RE, where the smoothest surfaces have the highest mean RE values. REs were not influenced by the low spore concentrations tested (3 x 10{sup -3} to 1.86 CFU/cm{sup 2}). The FNR data were consistent with RE data, showing a trend of smoother surfaces resulting in higher REs and lower FNRs. Stainless steel generally had the lowest mean FNR (0.123) and plastic had the highest mean FNR (0.479). The LOD{sub 90} varied with surface material, from 0.015 CFU/cm{sup 2} on stainless steel up to 0.039 on plastic. Selecting sampling locations on the basis of surface roughness and using roughness to interpret spore recovery data can improve sampling. Further, FNR values, calculated as a function of concentration and surface material, can be used pre-sampling to calculate the numbers of samples for statistical sampling plans with desired performance, and post-sampling to calculate the confidence in characterization and clearance decisions.

Joint Sandia/NIOSH exercise on aerosol contamination using the BROOM tool.

In February of 2005, a joint exercise involving Sandia National Laboratories (SNL) and the National Institute for Occupational Safety and Health (NIOSH) was conducted in Albuquerque, NM. The SNL participants included the team developing the Building Restoration Operations and Optimization Model (BROOM), a software product developed to expedite sampling and data management activities applicable to facility restoration following a biological contamination event. Integrated data-collection, data-management, and visualization software improve the efficiency of cleanup, minimize facility downtime, and provide a transparent basis for reopening. The exercise was held at an SNL facility, the Coronado Club, a now-closed social club for Sandia employees located on Kirtland Air Force Base. Both NIOSH and SNL had specific objectives for the exercise, and all objectives were met.

Investigation of a dual filter sampling method for gaseous and particulate fluoride

A two stage filter method designed to separate gaseous and particulate fluoride utilizing an alkaline treated filter to trap gaseous fluoride and post sampling heat treatment of the filters to promote desorption of gaseous fluoride from particulate phase was compared to standard impinger methods for the collection of hydrogen fluoride. Results indicate a high degree of comparability between the two methods. Initial studies involving the recovery of HF from dusted filters by heat treatment suggest recovery rates may vary directly with the amount of HF sampled for a given dust load.

Development of a Technology Roadmap for the Energy and Water Nexus

Energy and water are critical resources that are inextricably and reciprocally linked. The production of energy requires large volumes of water, and the treatment and distribution of water depends upon readily available, low-cost energy. For example, electricity production from thermoelectric power plants can use ∼140,000 million gallons of water per day for cooling—accounting for 39% of all freshwater withdrawals in the nation, second only to agriculture in the United States (Figure 1). Significant amounts of water are also needed for hydropower, extraction/refining of minerals for energy, and bio-fuel production. Electrical energy, on the other hand, is needed for water treatment (e.g., desalination, wastewater), pumping, and distribution. The amount of electricity used in water and wastewater industries is equivalent to the amount used in chemical, petroleum refining, and paper industries. These interdependencies, coupled with increasing demands for energy and diminishing availability of freshwater supplies, pose significant challenges to ensure the sustainability of these two critical resources. Examples of the interrelationships between energy and water use are shown in Figure 2.

Quick start users guide for the PATH/AWARE decision support system.

The Prioritization Analysis Tool for All-Hazards/Analyzer for Wide Area Restoration Effectiveness (PATH/AWARE) software system, developed by Sandia National Laboratories, is a comprehensive decision support tool designed to analyze situational awareness, as well as response and recovery actions, following a wide-area release of chemical, biological or radiological materials. The system provides capability to prioritize critical infrastructure assets and services for restoration. It also provides a capability to assess resource needs (e.g., number of sampling teams, laboratory capacity, decontamination units, etc.), timelines for consequence management activities, and costs. PATH/AWARE is a very comprehensive tool set with a considerable amount of database information managed through a Microsoft SQL (Structured Query Language) database engine, a Geographical Information System (GIS) engine that provides comprehensive mapping capabilities, as well as comprehensive decision logic to carry out the functional aspects of the tool set. This document covers the basic installation and operation of the PATH/AWARE tool in order to give the user enough information to start using the tool. A companion users manual is under development with greater specificity of the PATH/AWARE functionality.

A Simulation Tool for Assessing Sensor Performance in a Water Utility Contaminant Warning System

A set of software tools has been developed to enable performance testing of candidate on-line sensors in a utility-specific contaminant warning system design using simulation methods. Specific sensor performance characteristics such as noise, drift, detection level, and sampling rate can be input into the simulator and contamination scenarios can be run with a contaminant transport model such as EPANET using utility-specific pipe layouts. The sensor simulator is coupled with event detection software and together the two modules can provide an indication of how well the sensor will perform in a specific utility setting with user-defined contamination events. Water quality baseline data from utility measurements using on-line sensors are also incorporated into the analysis for a more realistic simulation. The software enables a determination of the rate of true positive, false positive, true negative, and false negative events during the simulation. Results from this analysis reveal that on-line residual chlorine and total organic carbon sensors combined with several different event detection algorithms can provide reliable detection for a variety of chemical contamination events. Study results further indicate this simulation tool concept is useful for pre-installation evaluation of contaminant warning system design in a specific utility setting.