Research Story of the Week

The Basics of Chemical and Biological Weapons Detectors

Photo
Detector in use.
[Src: Agilent Technologies]

Margaret E. Kosal, PhD

November 24, 2003


Introduction

Like modern canaries in a coal mine, the goal of chemical and biological weapons detectors and sensors is to alert to an imminent danger. This article's intent is to provide an overview of the technologies underlying detectors and the type of sensing systems currently employed or under near-term consideration for detecting chemical and biological warfare or terrorist weapons. The strategy for choosing a detector or a type of detector will also be explored - what are the comparative limitations and strengths of different detectors.

Current detector technology, for both chemical warfare (CW) and biological warfare (BW) agents, is strongest in terms of "detect to respond" or "detect to react" rather than "detect to warn." Most only respond when the threat is directly present. Alerting civilians, first responders or troops to the immediate danger of agent exposure is often the only goal of a detector. More sophisticated or additional instrumentation further refines the nature and concentration of the danger. Depending on the type of agent, the same technology is sometimes appropriate for microbial forensics or after-incident investigation, which is critical for law enforcement. A select few of these technologies come into play for on-site verification inspections as outlined in the Chemical Weapons Convention (CWC) or during UN-mandated inspections.

While many of the technologies discussed are appropriate for first-responders, this article is not meant to encompass diagnostic tools to determine medical or veterinary treatment after exposure or infection.

Biological Weapons Detectors

Biological agents come in many flavors - from delicate RNA-based filoviruses to robust spores of the Bacillus anthracis bacterium to toxins bestriding the margins of biological and chemical agents. These differences make the creation of a single detector for all biological agents challenging.

The "gold standard" for identification of microbiological species remains culturing - literally growing a colony of microbes on a nutrient containing surface (Petri dish) and observing it with the eye or through a microscope. Culturing is inexpensive and highly sensitive but slow. Roughly a minimum of a million (106) bacteria are necessary to form a visible colony. Detection of single cells is possible but only after long incubation times, usually days. Typical evaluation times are twelve to twenty-four hours for many bacteria but can exceed a week for exotic, slow-growing or more difficult to culture agents.

Remote or Standoff Detection

The initial criterion for monitoring and surveillance of potential biological agent at a distance is the observation of aerosolized masses (clouds). Spotting and evaluating the contents of a cloud is referred to as "standoff" detection. At a rudimentary level, these detector types aim to alert to the presence of an (approaching) cloud. Depending on the situation the recipient of that alert may be military, civil authorities, public health personnel or an individual. From that basic awareness, a more refined assessment of the contents, such as water droplets, inert inorganic material, dead biotic particulates or non-pathogenic microbes is pursued. Ideally a standoff detector will also be able to provide some information as to the nature of an aerosolized agent.

Cloud Recognition

One technique which is familiar from weather reporting is the use of Doppler radio detecting and reanging (radar). Using reflected radio waves, the shape, size, directionality and speed of a cloud can be monitored.[1] The elapsed time before the radio waves return to a receiver and the change in the radio waves' energy upon return to a receiver provide information about a cloud. For example, shape can offer clues to differentiate natural-occurring cumulus clouds from cigar-shaped ones (difficult to determine visually at night), which are indicative of aerosol release from a single source such as a plane or a moving vehicle.

Another tool for cloud detection and recognition, LIDAR, is based on the same physical principles as radar, except instead of bouncing longer wavelength radio waves off a target, higher energy light waves are used. An acronym for "Light Detection And Ranging," LIDAR is occasionally attributed to "Laser Identification and Ranging" by those who want to emphasize the recognition feature. Using lasers that generate light waves in the infrared, the ultraviolet and the visible portion of the electromagnetic spectrum, the multiple energy wavelengths of LIDAR furnish more detailed information, including three-dimensional imaging.[2] Limitations on detection distance and resolution are due to the collection and processing portions of the detector. The more specific the level of data desired, the closer the instruments must be located to the cloud.

Under controlled conditions, detection of aerosolized clouds at long distances has been achieved.[3] The temperature of a cloud can also be calculated using LIDAR data. Commercial applications for LIDAR include weather and upper atmosphere monitoring, elevation monitoring for planes and police monitoring of speeding automobiles. Water vapor and smog are potential interfering compounds for infrared-based LIDAR systems. The drawbacks are primarily financial and the current limited distance capability. LIDAR instruments are not cheap - costing about $4,000 for a simple LIDAR used for speed monitoring.

The U.S. Army's Long Range Biological Standoff Detection System (LR-BSDS) uses LIDAR-based technology to detect aerosol clouds from long distances. The Short Range Biological Standoff Detection System (SR-BSDS) combines infrared LIDAR with ultraviolet light reflectance (UV). The latter provides enhanced discrimination capabilities. Biological agents can be distinguished from non-biological material based on the excitation of the intracellular fluorescent compounds.[4] The most commonly targeted compounds are the amino acid tryptophan, the coenzyme nicotinamide adenine dinucleotide (NADH), the cellular energy storage molecule adenosine triphosphate (ATP) and the vitamin riboflavin. Identification of these compounds verifies that the sample is biological in origin. Possible false positives include pollen, molds, organic excreta and certain agricultural fertilizers based on decaying organic matter, e.g., "night soil."

Recent laboratory work using laser-induced breakdown spectroscopy (LIBS) has demonstrated the ability to remotely detect aerosolized and surface-adhered (on soil, rock, etc.) bacteria.[5,6] The LIBS-based systems not only detect the presence of an agent but also differentiate among bacterial species and among potential biological interferents (pollen, molds) with one instrument.

Point Detectors

Detectors that pass directly through or to which a potential biological agent-containing sample is introduced are referred to as "point" detectors. These require that the instrument, and usually the operator, physically enter a cloud and obtain a sample.

Aerosol Particle Sizers (APS)

Weaponized biological agents have characteristic physical dimensions. In order to be effective, agents must be small enough to not drop out of the cloud. Respirable particles have diameters between 0.5 and 20 µm (10-6 m). These are the particles which have the physiological potential to embed in the narrow passages (alveoli) or upper portions of the lung. Particles larger than 100 µm fall from the cloud; particles smaller than 0.5 µm are easily respired and do not remain in the lungs. Aerosol particle sizers (APS) take advantage of those size characteristics for detecting BW agents. A strongly uniform particle distribution in the size range associated with an inhalable risk or a substantial increase in numbers relative to a typical background may be indicative of the release of a biological agent. At the heart of APS instruments, nonetheless, is simply an attempt to detect higher than normal concentrations of airborne particles.

In APS systems, particles are drawn through an orifice into a steady high-speed air flow. The velocity of the carrier air remains constant throughout. The introduced particles accelerate at rates proportional to their size. Particles impact a collector or pass through a laser light beam to characterize the size.[7] While most particle sizers are fairly large and heavy systems, hand-held analyzers are commercially available.

Flow cytometry is a sophisticated particle counting technique in which particles are accelerated in a moving stream. Laser light scattering provides information with respect to the size, number and, when combined with fluorescent dye molecules, the chemistry of a sample. The ready combination of flow cytometry with UV or fluorescence methods provides more information about the nature of the material.[8]

Once the presence of aerosolized particles has been established, the next level of awareness relevant to detection of a potential BW threat is to seek specificity with regard to the agent, i.e., what exactly is it? These are sometimes referred to as discriminatory techniques. Detectors for agent identification primarily use two general ideas: (1) looking for a pathogen-specific tag or (2) taking the sample apart.

Immunoassays

Immunoassay-based detectors mimic the human body's natural immune system. The immune system produces highly specific proteins, called antibodies in response to antigens from foreign bacterium, toxins or other microbiological organisms. Antigens are molecules on the surface of the foreign microbes. Antibodies form strong and specific interactions with antigens. This specific response is the foundation of immunological detectors.

Disposable hand-held assay (HHA) test kits, such as enzyme-linked immunosorbent assays (ELISAs), or tickets for detecting biological warfare agents have been available since the early 1990s. Using laboratory-produced monoclonal antibodies, HHA tickets recognize the antigen in a sample to which that antibody would be produced if human infection occurred. This technique is pathogen-specific, i.e., one agent per test strip.

Immunoassys need some sort of optical signal generator - something that will "glow" when the detecting antibodies encounter a "hit." Typically, this is done with a fluorescent or chemiluminescent dye molecule that is chemically bonded to the detecting antibodies. The detection limit with fluorescence-based tags is on the order of 103 cells per mL and slightly lower for chemiluminescence.[9] The use of colloidal gold particles to generate a red indicator color without the need for a fluorescent light source has been used by the U.S. military and commercialized for the general public, although the detection limit is less sensitive than for other methods.

Some immunochromatographic tickets have exceedingly high reported false positive rates.[10] False positives are responses to something which the detector is not supposed to respond. A common false positive is response to a nonpathogenic "nearest neighbor" bacterial species found in the environment, i.e., mistaking Bacillus thuringiensis for B. anthracis. The flipside are false negatives, which are incidents in which an actual release is not detected. Such tests are both single use and respond to a single pathogen.

Among the disposable test strips currently available on the market, individual prices average approximately $20. While requiring less than twenty minutes for analysis and being easy-to-use, the antigen-antibody binding based tests are not sensitive. Illustratively, one of the best commercially available test strips for Bacillus anthracis requires greater than 10,000 spores for a positive reading, which is above the number the U.S. Army Medical Research Institute of Infectious Diseases (USAMRIID) cites as necessary to cause infection.[11] Additionally, cross-reaction with non-virulent related species generating false positives is frequently cited as a leading problem. Interfacing with a portable test strip reader permits more rapid analysis (less than five minutes) and simultaneous screening against multiple (greater than eight) agents. Multiplexed immunoassays are being developed to detect multiple pathogens.[12]

Another approach involves combing immunoassays with flow cytometry.[13] Individual microspheres or microfibers are labeled with both a specific antigen and multiple color-coded dyes to provide semi-quantitative assessment of multiple biological agents collected in a single sample.[14]

The Biological Detector (BD) portion of the U.S. military's Biological Integrated Detection System (BIDS) includes immunoassay-based sensors as part of its suite of detectors. Providing immunoassay tests for ten BW agents, including B. anthracis, Yersinia pestis, botulinum toxin A and staphylococcal enterotoxin B (SEB), the portable system requires substantial power, reagents, warm-up time, and is portable (135 lbs) only as far as the generator-carrying vehicle on which it is mounted can travel. Complete disclosure of the agents detected by BIDS has not been made available for security reasons.

Genetic Detection

In genetic-based detectors, DNA or RNA isolated from a sample is exposed to nucleic acid sequences, or oligonucleotides, which correspond to a suspected biological agent. These sequences are commonly referred to as "probes," as one can imagine a sequence "probing" a sample, seeking its genetic match. Similar to antibodies in immunoassay tests, these specific pieces of genetic material are typically tagged with an optical signaling molecule in order to indicate a positive result.

It is critical that probe sequences - the region of DNA or RNA targeted - be chosen well. If overly specific, a genuinely pathogenic strain may be missed yielding a false negative. Concurrently, if the chosen sequence is widely shared among a species or genus, it has the potential to respond to vaccine strains or to nearest-neighbor species, leading to false positives for innocuous non-pathogenic microbiologicals. A wise approach is to use oligonucleotides that target the virulence encoding DNA portion. In this way, genetically engineered species may also be identified.[15] Simple genetic-based ticket detectors are pathogen-specific, like the immunoassay counterparts.

Genetic-based detection is typically combined with an amplification technique, such as polymerase chain reaction (PCR) in order to generate larger quantities of genetic material in a shorter time frame than if the material were cultured.[16,17] Although many traditional instruments require a minimum of two to four hours, significant breakthroughs in thermocycling and microfluidics have led to reported analysis times of less than ten minutes. The amplified DNA can subsequently be compared to a library of unique oligonucleotides in order to identify the pathogen.

PCR and other DNA amplification techniques, while extremely powerful, are not without drawbacks. They are labor intensive, require consumable reagents, are restricted to liquid samples, offer marginal portability (typically exceeding 50 lbs), are demanding on power resources and are expensive. Different sample preparations are required for hardy anthrax spores than for a comparatively delicate filovirus. Currently, there is also a minimum of thirty minutes for protocol optimization.

Leading commercial manufacturers are advertising portable and handheld devices that combine PCR with genetic-based detection. While having significant advantageous in terms of specificity and detection limits over immunoassays, each suffers from limitations. Drawbacks that affect this type of system are the critical need for proper preparation, including thermal cycling for amplification, auxiliary reagents, high costs and highly trained operators of the devices. It is crucial that the pathogen sample be clean in order to differentiate from benign biological material in the environment. Nucleic acid probes also have finite life spans and generally require controlled storage conditions (e.g., freezers).

DNA microarrays or "chips" are being investigated for biological agent detection application. Allowing for parallel exposure of the potential pathogen to hundreds of specific substrate-immobilized oligonucleotides, these detection systems have significant potential.[18,19]

While immunoassays are limited by typically being single agent specific, the biggest liability for nucleic acid-based detectors is susceptibility to interferents. Isolation and purification of the sample are critical. Occasionally overlooked or underplayed in the excitement of innovative instrumentation is the criticality of sampling in BW agent detection. The way a sample is obtained and how it is handled can significantly affect the result, especially toward minimization of false positive responses. In addition to extricating biological agents from the surrounding environment, concentration of a sample can greatly enhance the ability to identify agents that are dilutely dispersed (but still of high enough concentration to have a deleterious effect on exposed humans).

Mass Spectrometry

The ability to characterize potential BW agents has been further enhanced by the use of mass spectrometry (MS) instruments. With this type of detection system, the sample of interest is fragmented into progressively smaller charged pieces ending with constituent amino acid or protein pieces. The charged fragments have different masses permitting physical differentiation. Furthermore the chemical groups yield characteristic fragment patterns - like fingerprints. While it is possible to solve small molecule fragmentation patterns manually, fingerprint libraries for comparison with known patterns are requisite for large molecules such as biologicals. Simple algorithms can provide reasonable "guesses," with estimations of uncertainty. Another approach uses mass spectroscopy to monitor the production of unique enzymatic metabolites generated by bacteria, fungi or rickettsia; the latter approach does not apply to viruses.

MS requires the separation of mixed samples, which is typically accomplished by subjecting the sample to a chromatographic separation prior to injection into the mass spectrometer. Chromatography refers to the separation of a mixture based on the component's physical interaction with a surface - typically the internal surface of a long column through which the mixture is pushed by a gas (GC) or a liquid (LC). The addition of chromatography significantly enhances the functionality of many detectors. Chromatography technically is not a means of detection, instead it is a means of separation. A detector is placed at the end of a chromatograph to analyze the outcome. GC can be used with either gaseous or liquid state samples. Tandem mass spectrometry, two sequential mass spectroscopy runs, is another method to separate sample mixtures, although less routine. The first MS run is used for separating the parts of a sample; the second MS run is used for detection.

GC-MS can detect, identify and differentiate diverse agents - bacteria, toxins and viruses.[20,21] For example, analysis of Yersinia species pathogens has been demonstrated.[22-24] In other works, the volatile biomarkers of B. anthracis and a nearest-neighbor species, Bacillus cereus, have been identified and differentiated using GC-MS and MS-MS techniques. Pyrolysis GC-MS, in which the sample is heated to a gas but not burned, can provide rapid analysis of intact, complete microbiologicals.[25,26] It is a comparatively harsh technique and not suited for detection of volatile biomarkers.

Commercial gas chromatography services, GC instruments and GC databases specific to detection of biological agents are available. Although exquisitely sensitive, current GC instruments are nevertheless bulky (and therefore, usually, remote from initial response sites), expensive ($30,000 to $150,000), require sample preparation and associated reagents and necessitate trained laboratory personnel.

Surface Acoustical Wave Sensors

Surface Acoustical Wave (SAW) systems are based on piezoelectric materials (those that produce an electrical current when subjected to pressure or mechanical stress) coated with antibodies or complimentary nucleic acid sequences. Binding of the target material causes a change in the mass of the piezoelectric sensing crystals which in turn changes the frequency at which the crystal vibrates under an electric current. This change in frequency is measured and alerts to the presence and, perhaps, the identity of the BW agent.[27,28] Sensitivities on the order of 105 - 106 cells have been reported.

Biowatch

In July 2003, the Department of Homeland Security working with the Environmental Protection Agency and the Centers for Disease Control revealed a thirty-one city program to monitor for BW agents in the air.[29] Dubbed "Biowatch," the system employs approximately five hundred air filters that are collected every twelve hours, and the filter contents are analyzed for BW agents using genetic-based detection equipment. This is an example of an attempt, on some level, to create a horizontal sensor web in which a single detector technology is distributed spatially across the country.

Chemical Weapons Detectors

Cutting-edge chemical techniques readily allow for the detection of single molecules.[30,31] Nevertheless, the experimental apparatus and conditions for such resolution are limited to sophisticated research laboratories. Detectors for chemical warfare agents and chemical terrorist weapons must function in demanding, real-world environments where price, portability and time are obligatory factors. Many CW agent detectors rely on adaptation of classical techniques from analytical chemistry to meet these demands.

As is the case for detecting biological agents, the most challenging aspect for chemical agent identification is often extracting the agent of interest from the other chemicals in the environment.

A brief note on terms: Vapor is the component of a liquid or a solid that has escaped the bulk phase, evaporated, and can be thought of as being in the gaseous state. Humidity is an example of liquid water in the vapor phase. While the ambient temperature is below the boiling point of water (212°F or 100°C) , some of the liquid is no longer readily in the bulk liquid phase. It's become a vapor or humidity. At standard ambient temperature (77°F or 25°C) and pressure, there are different CW agents that exist in each of the three physical states. Chlorine and phosgene are gases. Hydrogen cyanide and sarin are liquids. The vapor pressure - a measure of the comparative evaporation - is much greater for hydrogen cyanide than for sarin. Vapor pressure of liquids also contributes to agent persistency. Sarin is more persistant than hydrogen cyanide, but VX with a vapor pressure similar to motor oil is a more persistent liquid than either sarin or hydrogen cyanide. Riot control agents, such as chloroacetophenone (CN, "Mace") and 3-quinuclidinyl benzilate (BZ) are solids. The vapor phase of CN is readily apparent by its characteristic fragrant odor; whereas the crystalline solid BZ has negligible vapor pressure and no characteristic odor.

Remote or Standoff

Infrared Spectroscopy

Characteristic vibrational wavelengths of most CW agents occur in the infrared (IR) region of the electromagnetic spectrum. When IR light passes through a gas or vapor cloud, certain wavelengths of light are absorbed based on the chemical structure of the compounds in that cloud. Routine IR instruments measure the amount of light absorbed at a specific wavelength to look for a characteristic chemical group, such as the phosphorus-oxygen bond of nerve agents. More sophisticated instruments scan regions of the IR spectrum to generate a "fingerprint" pattern for individual chemicals. The corresponding distinguishing wavelengths are easily determined in a laboratory setting. With that data, huge libraries for comparison can be easily stored in portable instruments. Currently available IR spectrometers offer a limited level of standoff detection. The U.S. military's M21 Remote Sensing Chemical Agent Alarm (RSCAAL) employs infrared spectroscopy for standoff detection. Major limitations of IR-based sensors are cost, complexity and size of instrumentation.

Raman Spectroscopy

A technique similar to infrared spectroscopy, Raman spectroscopy also relies on known wavelengths of light at which organic molecules vibrate. Raman spectroscopy has also been used for non-destructive evaluation of CW agents in glass ampoules and bottles.[32] Raman is not applicable for identification of agents in munitions, as the technique requires a glass window through which light can pass.

Point Detectors

Colorimetric

Based on a visible color change, colorimetric indicators are the fastest, cheapest, lightest and easiest type of detector to use. The U.S. military employs two paper types, commonly referred to as M8 and M9 detection paper. The M8 paper detects and differentiates V-type nerve agents (VX), G-type nerve agents (sarin, soman) and H-type vesicants (sulfur mustard). The beige paper containing two dyes and an acid-base (pH) indicator changes to yellow when in contact with liquid sarin, green when in contact with liquid VX and red when in contact with a liquid mustard agent. Providing even a more general response, M9 paper does not differentiate among the agent types. Neither M8 nor M9 paper detects chemical agents in the vapor form - liquid must contact the paper surface for it to be a meaningful indicator. The papers are highly subject to false positives from bug spray, smoke, acetone, gasoline and strong bleach.

Unlike the simple color-changing of a dye on paper, colorimetric detection tubes provide a semi-quantitative indication of the amount of agent present. The tubes, which monitor one gas (or analyte) per test tube, are applicable to both vapors and gases. A typical system connects four or five tubes (less than a foot in length) to a small pump which pulls a vapor or gas sample through the tubes at a constant rate. Results are dependent on the analyte being tested, concentration and flow rate, but the tubes generally respond within a few minutes. Colorimetric detector tubes have been utilized for years by HazMat teams to identify industrial vapors and other noxious gases. They are both familiar to the first responder community and easy to use.

Electrochemical

In electrochemical or chemiresistor detectors, an electrical current changes in response to an interaction with a CW agent. The most common basis for an electrochemical gas sensor is a conducting wire or filament that is coated with a reactive material that oxidizes rapidly when it encounters a CW agent. The oxidation of the surface material exposes the conducting wire to air and the electrical resistance increases substantially. The change in current or increase in temperature is the signal for CW exposure. Other electrochemical detectors employ chemically selective membranes allowing only certain chemical types to pass - those which are required to complete a circuit. Again the change in current is the signal for CW presence. A newer type of chemiresistor instrument involves a quartz or silicon substrate which is coated with a conducting polymer. The degree of current change is dependent on the chemistry of the absorbing agent. The polymers provide limited specificity such that classes of CW agents can be differentiated. The response time for electrochemical sensors is generally very fast (less than a minute, often seconds.)

Electrochemical sensors are specific to single agents or, more commonly, to classes of analytes. Arrays of different sensors can be used to provide coverage for multiple types of agents.

Ion Mobility Spectrometry

Ion mobility spectrometry (IMS) or plasma chromatography relies on small differences in the velocity of ions along a cylindrical tube, a "drift tube", across which a constant electric field is applied.[33] Drift tubes have been miniaturized to the size of a credit card while retaining resolution. IMS instruments are quantitatively capable of detecting and identifying vapor-phase chemical agents and degradation products.[34] The response time is proportional to agent concentration; at "medium" to "high" ambient concentrations, response time is generally less than sixty seconds. Prominent examples of detectors using ionic mobility spectrometry include the U.S. military's Chemical Agent Monitor (CAM and ICAM) and Automatic Chemical Agent Alarm (ACADA). Airports frequently use IMS instruments for detecting explosives.

Mass Spectroscopy

The most sensitive and most reliable technique is mass spectrometry combined with gas chromatography (GC-MS). Specific CW agents, precursors, production by-products and degradation products are detectable and identifiable via GC-MS.[35-38] This type of detector also provides information regarding the concentration of CW agents.

All chemical agent detectors, save those incorporating chromatography, are limited when encountering mixtures. Commercial gas chromatography services, GC-MS instruments and GC-MS databases specific to detection of CW agents are available. Many combine standard commercial gas chromatography systems with proprietary databases. Although exquisitely sensitive, the average instrument is bulky (and therefore remote from initial response sites), expensive, requires sample preparation and associated reagents and needs technically trained personnel. GC-MS is the only CWC approved technique for on-site analysis during a challenge inspection.[39]

Flame Photometry

In flame photometric detection (FPD), a sample is ignited in a (very small) hydrogen flame. A characteristic emission spectrum is produced that serves as a fingerprint for the atoms in the compounds analyzed. In this way, a quantitative reading of the amount of a certain element, such as phosphorous or sulfur, in a sample can be detected. Optical filters can be selected for specificity of a target agent. A light detecting element (typically a photodiode) recognizes patterns that correspond to CW agents. An FPD detector can also be combined with a GC to improve complex mixture separation. Shortcomings include high cost and limited resolution compared to GC-MS. The French AP2C monitor and the Israeli CHASE detector use FPD technology.

Photoionization

Photoionization detector (PID) systems use ultraviolet (UV) light to ionize (remove the most loosely held electrons) from a vapor or gas. A detector measures the amount of ions based on a change in electrical current. PID systems are highly quantitative when compared to a calibrated known sample and provide excellent sensitivity in such situations. While popular, PID systems have very limited specificity and are highly subject to false positives in unknown or mixed environments. They are also costly and complex. Nonetheless, for applications such as leak testing, PIDs are appropriate.

Surface Acoustical Wave Sensors

Like the BW detector counterpart, SAW devices are based on piezoelectric materials that produce an electrical current when subjected to pressure or mechanical stress. Instead of antibodies or complimentary nucleic acid sequences, detectors for CW agents use individual piezoelectric quartz crystals (typically six or eight) or interdigitated electrodes coated with thin layers of different absorbent polymers.[40,41] A chemical will selectively absorb into the polymer based on chemical properties of each agent; generally each polymer-coated crystal will have an affinity for a different general class of organic vapors. The individual responses provide an array-based detection system. The innate sensitivity and response of SAW-based devices are limited by the polymer's absorption ability. Most SAW detectors incorporate an analytical preconcentrator in order to overcome these limitations; the commercially available SAW-based instrument incorporates a GC prior to exposure to the SAW detector. The SAW device alone is very small - the size of a penny. The U.S. military's Joint Chemical Agent Detector (JCAD) employs SAW-based technology. The JCAD is a handheld, lightweight CW detector. Reportedly, it is enabled to detect new forms of nerve agents.

The absorbent polymers used in SAW devices are susceptible to damage from certain highly reactive vapors. Hydrofluoric acid (HF) is one such vapor. HF is also a degradation product from the hydrolysis (chemical break down due to water or ambient humidity exposure) of sarin, soman and cyclosarin. The polymers used are often susceptible to interference from absorption of water and, therefore, the sensors must be calibrated (and re-calibrated) to account for ambient relative humidity.

Enzyme-Based

Enzyme or immunoassays approaches have been utilized for military and commercial CW detectors. Some enzyme-based CW detection systems exploit the intent of organophosphate nerve agents to bind to acetylcholinesterase - an enzyme - as a detection technique. Enzyme-linked immunoassays (ELISAs) have been developed, much like the BW agent counterpart, with specificity for G-type nerve agents.[42] Other systems, exploit the natural enzyme that catalytically hydrolyzes (breaks down in the presence of water) organophosphates. Organophosphorous hydrolase (OPH) can be incorporated into hand-held assays or tickets.[43] A pH sensitive probe reacts to change in acidity due to the hydrolysis of G-type nerve agents. The response can be as simple as a colorimetric pH indicator changing from red to blue or a potentiometric electrode.

Non-Destructive Evaluation

Isotopic Neutron Spectroscopy

Isotopic neutron spectroscopy was developed to provide information about an agent in a sealed container, for example, old munitions.[44] Using neutrons from controlled gamma radiation to atomically interrogate a container's contents, a detector records the energies of the neutron-atom interactions. The magnitude of these interactions serves as a "signature" of the chemical elements inside. Signatures that correspond to high nitrogen contents are typical of high explosives. Those results which indicate significant phosphorous are indicative of nerve agents. High sulfur and chlorine readings are indicative of a mustard blister agent. Experimental results are compared with the signatures of known chemical compounds to identify the specific agents.

The Portable Isotopic Neutron Spectroscopy System (PINS) is employed in the field to differentiate traditional munitions from those containing CW agents. While readily "field-able" and exceeding any other system for verification of internal contents, the system's portability is marginal. PINS consists of five gym locker-sized boxes of instrumentation. Among the necessary expendables is liquid nitrogen. The set-up and break-down of the instrument consumes more time than the actual assay (fifteen minutes or less). Nonetheless, for verification of CW destruction purposes, PINS has the potential to be an extremely valuable and efficient tool. Not only can it detect the difference between conventional and CW-containing munitions, it can also identify munitions containing simulants, water, sand or concrete.
Acoustic Resonance Spectroscopy (ARS) and Swept Frequency Acoustic Interferometry (SFAI)

The two related techniques ARS and SFAI rely on the fundamental difference in the speed of sound through a solid versus a liquid. Detectors employing these techniques are used to classify munitions by content. ARS and SFAI monitor the change in magnitude of sound (resonance) vibrating through a material over a range (typically about 30kHz for ARS, larger frequency ranges for SFAI) of sound frequencies.[45] These are important for the determination of the contents of old CW munitions. If the fill on a specific munition type is unequivocally known (e.g., a 155mm shell containing thickened sarin) for a minimum number of samples (approximately six), ARS can record a "template" for that material and use it for direct comparisons with unknown munitions. SFAI is also appropriate for gases. The systems are man-portable, and each instrument run is complete within a few minutes. ARS and SFAI are also used for evaluation of drums containing nuclear (waste) material, integrity of seals on nuclear material, and integrity of building materials. They are being explored for medical imaging.

Validation and Standardization Issues

In this article, the intent has been to provide an overview of currently employed technologies for detection of chemical and biological agents. As new technologies emerge, there remains a need to insure third-party validation of the results and pursue significant real-world testing.[46,47] There is little doubt of the value in developing new and better instrumentation, much of which may arise from fundamental research. In a funding flurry and a ripe political climate, new devices and new experimental techniques need to be subject to extensive scrutiny and validation procedures. Among the foremost reasons are to limit false negatives and false positives at real-world sites. Excessive false positives can lead to response fatigue and ignoring a real incident; a false negative that fails to detect a CBW agent release could cause a disaster of the highest order - the loss of human life.

Conclusions

There currently exist a wide variety of techniques that provide excellent detection capabilities for CBW agents. Each, however, has drawbacks and limitations. The prospect of a single detector amenable to all CW and BW agents is laudable, although unrealistic with current technology. Layered detectors and sensors that function together in a web-like manner to monitor progressively more refined levels - from cloud and particle detection to differentiation between biological and nonbiological components to concentration information - are a near-term approach to unified and comprehensive CBW detection. This strategy involves the development of vertical sensor webs in which different levels of detection are optimized in addition to a horizontal sensor web (the same detector distributed spatially). An additional component should be intentional redundancies to limit false positives and false negatives. Integrating systems to synergistically operate will be a significant technical challenge as most devices have been designed and manufactured as stand-alone instruments. There is also a considerable political challenge in the design and implementation of such a sensor system.


References
1. Ember L, "From weather radars to chem-bio detectors," Chemical & Engineering News, 80, 2002, pp. 23-42.
2. Weibring P, Ember L, and Svanber S, "Versatile mobile lidar system for environmental monitoring," Applied Optics, 42, 2003, pp. 3583-3594.
3. Lee KJ, Youngsikpark, Bunkin A, Nunes R, Pershin S, and Voliak K, "Helicopter-based lidar system for monitoring the upper ocean and terrain surface," Applied Optics, 41, 2002, pp. 401-406.
4. Lognoli D, Lamenti G, Tirelli D, Tiano P, Tomaselli L, and Pantani L, "Detection and characterization of biodeteriogens on stone cultural heritage by fluorescence lidar," Applied Optics, 41, 2002, pp. 1780-1787.
5. Morel S, Leone N, Adam P, and Amourous J, "Detection of bacteria by time-resolved laser-induced breakdown spectroscopy," Applied Optics, 42, 2003, pp. 6184-6191.
6. Samuels AL, DeLucia KL, McNesby KL, and Miziolek A, "Laser-induced breakdown spectroscopy of bacterial spores, molds, pollens and protein: initial studies of discrimination potential," Applied Optics, 42, 2003, pp. 6205-6209.
7. Liu BYH, Yoo S-H, and Chase S, "Lower detection limit of aerosol particle counters," Journal of the Institute of Environmental Sciences, 38, 1995, pp. 31-37.
8. Ho J, "Future of biological aerosol detection," Analytica Chimica Acta, 457, 2002, pp. 125-148.
9. Iqbal SS, Mayo MW, Bruno JG, Bronk BV, Batt CA, and Chambers JP, "A review of molecular recognition technologies for detection of biological threats," Biosensors & Bioelectronics, 15, 2000, pp. 549-578.
10. Smithson AE and Levy L-A, "Ataxi: The Chemical and Biological Terrorism Threat and the US Response," , (The Henry L. Stimson Center; Washington, D.C.), 1999, p. 185.
11. Enserink M, "Biodefense hampered by inadequate tests," Science, 294, 2001, pp. 1266-1267.
12. McBride MT, Gammon S, M Pitesky, O'Brien TW, Smith T, Aldrich J, Langlois RG, Colston B, and Venkateswaran KS, "Multiplexed liquid arrays for simultaneous detection of simulants of biological warfare agents," Analytical Chemistry, 75, 2003, pp. 1924-1930.
13. Lisi PJ, Huang CW, and Hoffman RA, "A fluorescence immunoassay for soluble antigens employing flow cytometric detection," Clinica Chimica Acta, 120, 1982, pp. 171-179.
14. Park MK, Briles DE, and Nahm MH, "A latex bead-based flow cytometric immunoassay capable of simultaneous typing of multiple pneumococcal serotypes (Multibead assay)," Clinical and Diagnostic Laboratory Immunology, 7, 2000, pp. 486-489.
15. Slezak T, Kuczmarski T, Ott L, and Torres C, "Comparative genomic tools applied to bioterrorism defense," Briefs in Bioinformatics, 4, 2003, pp. 133-149.
16. Belgrader P, Benett W, Hadley D, Long G, Mariella R, Milanovich F, Nasarabadi S, Nelson W, Richards J, and Stratton P, "Rapid pathogen detection using a microchip PCR array instrument," Clinical Chemistry, 44, 1998, pp. 2191-2194.
17. Jones M, Alland D, Marras M, El-Hajj H, Taylor MT, and McMillan W, "Rapid and sensitive detection of mycobacterium DNA using Cepheid SmartCycler and Tube Lysis system," Clinical Chemistry, 47, 2001, p. 1917.
18. Cheng J, Frotina P, Surrey S, Kricka LJ, and Wilding P, "Microchip-based devices for molecular diagnosis of genetic diseases," Molecular Diagnostics, 1, 1996, pp. 183-200.
19. Du H, Miller BL, and Krauss TD, "Hybridization-based unquenching of DNA hairpins on Au surfaces: prototypical 'molecular beacon' biosensors," Journal of the American Chemical Society, 125, 2003, pp. 4012-4013.
20. Beverly MB, Voorhees KJ, Hadfield TL, and Cody RB, "Electron monochromator mass spectroscopy for the analysis of whole bacteria and bacterial spores," Analytical Chemistry, 72, 2000, pp. 2428-2432.
21. Fuerstenau SD, Benner WH, Thomas JJ, Brigidou C, Bothner B, and Siuzdak G, "Mass spectrometry of an intact virus," Angewandte Chemie, 40, 2001, pp. 542-544.
22. Fox A, Black GE, Fox K, and Rostovtseva S, "Determination of carbohydrate profiles of Bacillus anthracis and Bacillus cereus including identification of O-methyl methylpentoses by using gas chromatography-mass spectrometry," Journal of Clinical Microbiology, 31, 1993, pp. 887-894.
23. Jantzen E and Lassen J, "Characterization of Yersinia pestis species by analysis of whole-cell fatty acids," International Journal of System Bacteriology, 30, 1980, pp. 421-428.
24. Leclercq A, Wauters G, Decallonne J, El Lioui M, and Vivegnis J, "Usefulness of cellular fatty acid patterns for identification and pathogenicity of Yersinia species," Medical Microbiology Letters, 5, 1996, pp. 182-194.
25. Morgan CH, Mowry C, Manginell RP, Frye-Mason GC, Kottenstette RJ, and Lewis P, "Rapid identification of bacteria with miniaturized pyrolysis/GC analysis," Proceedings of SPIE-The International Society for Optical Engineering (Advanced Environmental and Chemical Sensing Technology, 4205, 2001, pp. 199-206.
26. Snyder AP, Maswadeh WM, Parsons JA, Tripathi A, Meuzelaar HLC, Dworzanski JP, and Kim MG, "Field detection of Bacillus spore aerosols with stand-alone pyrolysis-gas chromatography-ion mobility spectrometry," Field Analytical Chemistry Techniques, 3, 1999, pp. 315-326.
27. Grate JW, Martin SJ, and White RM, "Acoustic wave microsensors," Analytical Chemistry, 650, 1993, p. 940A.
28. Skladel P, "Piezoeletric quartz crystal sensors applied for bioanalytical assays and characterization of affinity interactions," Journal of the Brazilian Chemical Society, 14, 2003, pp. 491-502.
29. Mintz J, "U.S. provides a peek at air sensor program," The Washington Post, Nov. 15, 2003, p. A03.
30. Single-Molecule Detection in Solution Methods and Applications, Zander C, Enderlein J, and Keller RA, eds. (Hoboken, NJ: Wiley-VCH, 2002).
31. Single-Molecule Optical Detection, Imaging and Spectroscopy, Basche T, Moerner WE, Orrit M, and Wilding P, eds. (New York: John Wiley & Sons, 1996).
32. Christensen S, MacIver, B, Procell L, Sorrick D, Carrabba, M, and Bello J, "Nonintrusive analysis of chemical agent identification sets using a portable fiber optic raman spectrometer," Applied Spectroscopy, 53, 1999, pp. 850-855.
33. Eiceman GA and Karpas Z, Ion Mobility Spectrometry (Boca Raton, FL: CRC Press, 1994).
34. Steiner WE, Clowers BH, Matz LM, Siems WF, and Hill HH, "Rapid screening of aqueous chemical warfare agent degradation products: ambient pressure ion mobility mass spectrometry," Analytical Chemistry, 74, 2002, pp. 4342-4352.
35. Encyclopedia of Analytical Chemistry: Instrumentation and Applications: Chemical Warfare Agents Detection, Meyers RA, ed. (Chichester: John Wiley & Sons Ltd., 2000).
36. Occolowitz JL and White GL, "The mass spectrometry of esters of phosphorous and phosphonic acids," Analytical Chemistry, 35, 1963, pp. 1179-1182.
37. Driskell WJ, Shih M, Needham LL, and Barr DB, "Quantitation of organophosphorus nerve agent metabolites using isotope dilution gas chromatography-tandem mass spectrometry," Journal of Analytical Toxicology, 26, 2002, pp. 6-10.
38. Black RM, Clarke RJ, Read RW, and Reid MTJ, "Application of gas-chromatography-mass spectrometry and gas chromatography-tandem mass spectrometry to the analysis of chemical warfare agent, found to contain residues of the nerve agent sarin, suphur mustard and their degradation products," Journal of Chromatography A, 662, 1994, pp. 301-321.
39. Erickson B, "The Chemical Weapons Convention redefines 'analytical challenge'm," Analytical Chemistry, 70, 1998, pp. 397A-400A.
40. Niewenhuizen MS, Harteveld, and JLN, Sensors and Actuators B, 40, 1997, pp. 167-173.
41. Williams D and Pappas G, Field Analytical Chemistry Techniques, 3, 1999, pp. 45-53.
42. Lenz DE, Brimfield AA, and Cook LA, "Development of immunoassays for detection of chemical warfare agents," Immunochemical Technology for Environmental Applications, Aga DS and Thurman EM, eds. (Washington, DC: American Chemical Society, 1997), pp. 77-86.
43. Varfolomeyev S, Kurichkin I, Eremenko A, and Efremenko E, "Chemical and biological safety. Biosensors and nanotechnological methods for the detection and monitoring of chemical agents," Pure and Applied Chemistry, 74, 2002, pp. 2311-2316.
44. Parker WE, Buckley WM, Kreek SA, Caffrey AJ, Mauger GJ, Lavietes AD, and Dougan AD, "A portable system for nuclear, chemical agent and explosives identification," American Institute of Physics Conference Proceedings, 576(1), 2001, pp. 1073-1076.
45. Trujillo MT, Trujillo CC, Miller DA, and Baiardo J, "Acoustic resonance spectroscopy for structural evaluation," American Institute of Physics Conference Proceedings, 673(1), 2003, pp. 211-217.
46. Emanuel PA, Chue C, Kerr L, and Cullin D, "Validating the performance of biological detection equipment: the role of the federal government," Biosecurity and Bioterrorism, 1, 2003, pp. 131-137.
47. Turner RB, "Transitioning analytical instrumentation from the laboratory to harsh environments," Pure and Applied Chemistry, 74, 2002, pp. 2317-2322.


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Author(s): Margaret Kosal
Related Resources: Chem/Bio, Weekly Story
Date Created: November 24, 2003
Date Updated: November 25, 2003
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