Biological Detection Systems Research Articles

Plant Analysis 2008 – Planar Chromatography

For 70 years, planar chromatography has been closely connected with the analysis of plants. Also today in its most advanced form "high performance thin-layer chromatography" (HPTLC), the technique plays an important role in identification of medicinal plants and dietary supplements. Detection of adulteration and quantitative determination of marker substances are other widely used applications. The advancement of instrumentation and methodological concepts has created a basis for reliable qualitative and quantitative results in HPTLC. Remarkable progress has been made in the manageability of the entire planar chromatographic process, particularly in the control of chromatogram development and the utilization of images to obtain and compare information about separated samples. Integration of biological detection systems as well as hyphenation to mass spectrometry has widened the applicability of planar chromatography as an analytical technique that is both orthogonal and complementary to HPLC.

Lidars: A Key Component of Urban Biodefense

A biological aerosol attack in a city could infect tens of thousands of people. In the absence of announcements by the attacker or detection by present point detection systems, victims would be unaware of their exposure prior to developing symptoms. Since infections are most effectively countered before the onset of symptoms, detection technologies that provide early awareness of an attack should be given high priority. Current biological point detection (BPD) systems collect environmental air samples and then analyze them in laboratories so as to permit detection within 12-36 hours of an attack. Improvements in the pipeline may reduce this lag time to as little as a few hours. However, BPD systems have inherent weaknesses when used to detect and respond to an aerosol attack. The likelihood of a limited number of BPD systems intercepting an aerosol plume in a vast attack space may be low. Moreover, BPD systems do not provide critical information needed for response, such as the source location, precise time, and geographic reach of an attack. The missing information would invaluably guide prophylaxis distribution, identification of contaminated areas, and criminal apprehension. This article describes how lidars used for real-time observation of aerosol plumes could complement BPD systems by providing fine-scale spatial and temporal information. A lidar system also could be used to corroborate positive BPD system results, to improve reaction to positive results, and/or to provide an independent basis for low-regret protective steps. Lidar systems can resolve key biodefense challenges, and this article describes three compatible concepts of operations. Leveraging lessons from a lidar system now operating at the Pentagon, a test of an expanded lidar network would provide immediate protection for key Washington, DC, assets, demonstrate the synergy of BPD systems and lidars, and provide a test bed for research to improve lidar's shortcomings.

Detection of nitric oxide in single cells

Nitric oxide (NO) is endogenously generated by nitric oxide synthase (NOS) enzymes and is involved in a surprisingly wide range of biological functions. As efforts are made to elucidate the regulatory mechanisms of NOS expression and function, there is increasing interest in following NOS activity directly by monitoring NO production. Additionally, spatial and temporal measurements of NO are important for understanding its function and metabolism. In this work, developments in technology enabling NO detection in biological systems are reviewed. Measuring NO at single cell levels is important as NOS is heterogeneously distributed; however, such measurements are difficult as physiological NO levels are in the low nanomolar to low micromolar range. Here, three categories of analytical techniques enabling NO detection at single cell levels are highlighted: fluorescence microscopy, capillary electrophoresis with laser induced fluorescence detection, and electrochemistry. For each, the basic principles, performance, applications, figures of merits and limitations are presented in terms of single cell NO detection.

Rationally designed nanostructures for surface-enhanced Raman spectroscopy.

Research on surface-enhanced Raman spectroscopy (SERS) is an area of intense interest because the technique allows one to probe small collections of, and in certain cases, individual molecules using relatively straightforward spectroscopic techniques and nanostructured substrates. Researchers in this area have attempted to develop many new technological innovations including high sensitivity chemical and biological detection systems, labeling schemes for authentication and tracking purposes, and dual scanning-probe/spectroscopic techniques that simultaneously provide topographical and spectroscopic information about an underlying surface or nanostructure. However, progress has been hampered by the inability of researchers to fabricate substrates with the high sensitivity, tunability, robustness, and reproducibility necessary for truly practical and successful SERS-based systems. These limitations have been due in part to a relative lack of control over the nanoscale features of Raman substrates that are responsible for the enhancement. With the advent of nanotechnology, new approaches are being developed to overcome these issues and produce substrates with higher sensitivity, stability, and reproducibility. This tutorial review focuses on recent progress in the design and fabrication of substrates for surface-enhanced Raman spectroscopy, with an emphasis on the influence of nanotechnology.

Noise considerations in field-effect biosensors

Field-effect sensors used to detect and identify biological species have been proposed as alternatives to other methods such as fluorescence deoxyribonucleic acid (DNA) microarrays. Sensors fabricated using commercial complementary metal-oxide-semiconductor technology would enable low-cost and highly integrated biological detection systems. In this paper, the small-signal and noise modeling of biosensors implemented with electrolyte-insulator-semiconductor structures is studied, with emphasis on design guidelines for low-noise performance. In doing so, a modified form of the general charge sheet metal-oxide-semiconductor field-effect transistor model that better fits the electrolyte-insulator-semiconductor structure is used. It is discussed how if the reference electrode and the insulator-electrolyte generate no noise associated with charge transport, then the main noise mechanisms are the resistive losses of the electrolyte and the low-frequency noise of the field-effect transistor. It is also found that for realistic sensor geometries and high electrolyte concentrations, the noise from the field-effect transistor (FET) dominates the thermal noise from the electrolyte resistance, and the optimal biasing point for the FET for minimum noise is found to be around moderate inversion.

Synthesis of a hydrophilic and non-ionic anthracene derivative, the N, N′-di-(2,3-dihydroxypropyl)-9,10-anthracenedipropanamide as a chemical trap for singlet molecular oxygen detection in biological systems

We report herein the synthesis of a new hydrophilic and non-ionic anthracene derivative, the N, N′-di-(2,3-dihydroxypropyl)-9,10-anthracenedipropanamide. The evaluation of this compound as chemical trap of singlet molecular oxygen by using labeling experiments and HPLC–MS analysis showed that it could be efficiently used in biological investigations.

Preparation and evaluation of spore-specific affinity-augmented bio-imprinted beads

A novel, affinity-augmented, bacterial spore-imprinted, bead material was synthesized, based on a procedure developed for vegetative bacteria. The imprinted beads were intended as a front-end spore capture/concentration stage of an integrated biological detection system. Our approach involved embedding bead surfaces with Bacillus thuringiensis kurstaki (Bt) spores (as a surrogate for Bacillus anthracis) during synthesis. Subsequent steps involved lithographic deactivation using a perfluoroether; spore removal to create imprint sites; and coating imprints with the lectin, concanavalin A, to provide general affinity. The synthesis of the intended material with the desired imprints was verified by scanning electron and confocal laser-scanning microscopy. The material was evaluated using spore-binding assays with either Bt or Bacillus subtilis (Bs) spores. The binding assays indicated strong spore-binding capability and a robust imprinting effect that accounted for 25% additional binding over non-imprinted controls. The binding assay results also indicated that further refinement of the surface deactivation procedure would enhance the performance of the imprinted substrate.

Capture of airborne nanoparticles in swirling flows using non-uniform electrostatic fields for bio-sensor applications

Collection of biological particles is the first and critical step for any biological agent detection system. Towards our goal of capturing and detecting airborne biological entities in real time, here we investigate on the design of an electrostatic particle capture system. We report on the capture of airborne 100 nm diameter polystyrene nanoparticles as a model system, in swirling flows under non-uniform electrostatic fields with an electrospray aerosol generator and a homemade particle collector. The particle collector has five positive electrodes on the bottom and one large grounded electrode on the top. The nanoparticles coming into the collector were slowed down during their swirling and stayed in the collector long before leaving the collector. Silicon chips were placed on the bottom electrodes and the electrostatically captured particles were counted as a function of flow rates, electrode positions, bias voltages, and capture times by epifluorescent images and scanning electron micrographs (SEMs). Particles captured in the electrode at the center of the collector were much less than those on the surrounding four electrodes and 10–25% of the particles with negative charges entering the collector were captured on the bottom electrodes at a flow rate of 1.1 l/min and an applied potential of 2 kV. Particle capture increased with decreasing flow rates. We also simulated flow and electrical fields separately, and found the positional trends to be in good agreement with the measurements. This collector is well adaptable to integration with micro resonator devices and can be used for real-time monitoring of bioaerosols.

Editorial [Hot Topic: Lipid-Based Nanosystems and Complexes in Experimental and Clinical Therapeutics (Guest Editors: A. Agrawal and S.M. Moghimi)

An intriguing spectrum of novel nanoparticulate systems and complex nanostructures, some with remarkable diagnostic and biomimetic properties and others suitable for site-specific drug delivery and targeting are in existence today. Examples include quantum dots, capable of labelling biological systems for detection by optical or electrical means, tris-malonic acid derivatives of the fullerene C60, which exhibit superoxide dismutase mimetic properties, and porous titanium oxide and silicon nanoparticulate systems with sustained drug release properties. The ability to create these assemblies as well as other unusual nanostructures such as bundles, tubes, and sheets holds promise for new and powerful diagnostic and drug delivery systems, which will surely change the foundations of disease treatment and diagnosis. These technological innovations and nanoscience approaches to particle design are the focus of the US National Institute of Health's Nanomedicine Roadmap Initiative and the National Cancer Institute's Alliance for Nanotechnology in Cancer and several new European Commission-funded initiatives. Indeed, February 2004 saw the birth of Nano2Life, Europe's first Network of Excellence in nanobiotechnology, and in February 2005, the European Science Foundation launched its Scientific Forward Look on Nanomedicine Initiative, with strong emphasis on engineering of smart nanoparticle-based medicines and nanodevices. Although within the remit of the above mentioned nanoscience initiatives, lipid-based nanosystems such as nanoemulsions, lipid-core micelles, small unilamellar vesicles and variations thereof, have long been in existence and some have long been improving patient's lives. Indeed, lipid-based nanoformulations are among the most attractive candidates for improving drug solubility and for site-specific targeting following parenteral administration. This Theme Issue of Current Drug Delivery features contributions focusing on selected molecularbased strategies and nanoscience approaches to design, development, and site-specific targeting of a wide range of lipid-based nanocarriers and complex systems currently at preclinical and clinical stages. Specifically, great strides are being made with such complexes and nanosystems in combating the growth and spread of cancerous tissues (e.g., through exploitation of angiogenic tumour vasculature, combination chemotherapy, and endogenous triggered activation and release of encapsulated lipid pro-drugs), treatment of macrophage infections (through exploitation of macrophage clearance mechanisms), gene transfer (by breaching the endo-lysosomal barrier with cationic lipid vectors) and stimulation of immune responses to antigens (with the aid of vesicular systems and lipid-complexes with self-adjuvanting properties), and are discussed in the first thirteen articles in this Theme Issue. Although, lipidbased nanocarriers may overcome solubility or stability issues for the drug and minimize drug-induced side effects through favourable pharmacokinetic profiles and site-specific targeting, there are significant toxicity issues with carriers themselves that need to be addressed. Selected toxicity issues are dealt with in the last two articles. The first contribution discusses toxicogenomics of the delivery system and the impact of the microarray technology; in assessing cellular toxicity, while in the final article, the reader's attention is drawn towards pseudoallergic reactions, which are believed to be secondary to complement activation. Here, endogenous lipid-based delivery vehicles (lipoproteins) seem to play a complex modulatory role in complement-mediated pseudoallergic responses to nanomedicines. We believe that this Issue has provided a broad sample of the state of the art. We are indebted to all contributors for their attempt to communicate and discuss the most promising ideas, approaches, applications and developments of the most advanced lipid-based nanomedicines.

If You Build It, Will They Come?

Tom Petzinger, cofounder of Launchcyte, a for-profit Pittsburgh biotech incubator now located in the Greenhouse, is a former business writer for the Wall Street Journal. Four years ago, he and colleagues started Launchcyte with ten people and dedicated facilities.“The incubator’s primary objective is creating value for private investors,” says Petzinger. “If in doing so, we serve a regional economic development objective, that is great. But the latter is not the principal objective. An opposite scenario could work so long as the secondary objective [i.e., creating value for the investors] is treated with respect. There is no point in wasting common resources on businesses that have no viability.”“In the early stage of development, the technologies that are being commercialized are premature, and the risks are high,” Petzinger says. “That has been the case in everything we have done. It is not easy to find an investor to shoulder the burden of technology risks as investors would rather shoulder the burden of market risk and competitive risk. This is where organizations like the Greenhouse or the Michigan Initiative can really fill an important gap.”According to Petzinger, the business model for a biotech startup should be focused on accumulating intellectual property and then harvesting value in an exit, either by licensing intellectual property or selling it to a pharmaceutical company, which would then undertake the more expensive clinical trials and FDA approvals. It is far less likely to build a biotech business model based on revenue.To date, none of the original companies formed at Launchcyte have folded, but the business model has changed. “We ceased being an incubator,” says Petzinger. Launchcyte’s staff has shrunk to Petzinger and an office manager. “I learned that what is most important is selecting technologies appropriately, putting in the right teams, and sending them to market.” At the end of 2003, Launchcyte received a $1 million cash infusion from the University of Pittsburgh, University of Pittsburgh Medical Center, and private investors. Launchcyte is essentially functioning now as a parallel entrepreneur in pursuit of multiple opportunities simultaneously. It is more a venture capital fund “but heavy on the venture and light on the capital,” Petzinger quips. Launchcyte has invested around $12 million dollars in its original five companies; currently a sixth one is being formed. “Every one of the companies drives its market,” Petzinger says. “Three of them have revenues, real money from the marketplace. That is the best source of financing in the world.”Petzinger remembers that when he was working in Allentown, Pennsylvania and the steel mills went away, local officials considered creating jobs by luring in, among other industries, a dirigible factory. “Zeppelins, like the Hindenburg,” Petzinger recalls. “This is better.”

Polydiacetylene (PDA)-based colorimetric detection of biotin–streptavidin interactions

The natural binding of streptavidin (STA) for a small molecule, biotin, has made it a useful tool in specific targeting application, due to their most specific noncovalent biological interactions in nature. Here, we describe a polydiacetylene (PDA)-based colorimetric biosensor to detect the STA-biotin interactions. We synthesized two kinds of biotin-labeled 10,12-pentacosadiynoic acid (PCDA) monomers having 2(ethylene oxide) or 3(ethylene oxide) spacer and prepared biotin-modified PDA liposomes using them. Upon the addition of STA, the modified PDA liposomes showed a color change from blue to red in about 1h followed by their aggregation and precipitation. The aggregation was found to be due to the cross-linking between the PDA liposomes caused by the STA having four biotin binding sites, which was confirmed by their TEM analysis. Since the STA-biotin interactions are widely adopted for various biological detection systems; the biotin-modified PDA sensor developed in this study has enormous potentials for the development of various colorimetric biosensors.

Vapor phase detection of a narcotic using surface acoustic wave immunoassay sensors

Currently, the narcotic sniffing dog remains the most accurate, reliable, and widely used sensing technology in the war on drugs. However, recent studies done at the Institute for Biological Detection Systems at Auburn University, Auburn, AL, have shown that in the presence of extraneous odors (nontarget odors), these animals show a higher propensity for so-called false alarms. For this reason, there have been an increasing demand for a portable, highly specific vapor-sensing device capable of distinguishing a target vapor signature in a complex odor. In this paper, we present the results of a series of experiments demonstrating real-time vapor phase detection of cocaine molecules. A distinctive response or signature was observed under laboratory conditions, where the cocaine vapors were presented using an INEL vapor generator and under "field" conditions facilitated by the Georgia Bureau of Investigation Crime Lab. For these experiments, the sensor component was an ST-X quartz resonator with a center frequency of approximately 250-MHz. Anti-benzoylecgonine (anti-BZE) antibodies are attached to the electrodes on the device surface via a protein-A cross linker. We observed a large transient frequency shift accompanied by baseline shift with the anti-BZE coated sensor. After repeated experiments and the use of numerous controls, we believe that we have achieved real-time molecular recognition of cocaine molecules.

Regional and whole-body markers of nitric oxide production following hyperemic stimuli

The measurement of nitric oxide (NO) bioavailability is of great clinical interest in the assessment of vascular health. However, NO is rapidly oxidized to form nitrite and nitrate and thus its direct detection in biological systems is difficult. Venous plasma nitrite (nM concentrations) has been shown to be a marker of forearm NO production following pharmacological stimulation of the endothelium utilizing acetylcholine (Ach). In the present study, we demonstrate, within 15 apparently healthy subjects (34.1 ± 7.3 years), that reactive hyperemia of the forearm, a physiological endothelial stimulus, results in a 52.5% increase in mean plasma nitrite concentrations (415 ± 64.0 to 634 ± 57.1 nM, P = 0.015). However, plasma nitrite is readily oxidized to nitrate within plasma, and thus its utility as a marker of NO production within the clinical setting may be limited. Alternatively, NOx (predominantly nitrate) is relatively stable in plasma (μM concentrations), but is produced by sources other than the vasculature and has been shown to be unsuitable as a measure of localized NO production. We reasoned that the principle source of NOx generation during exercise is NO production and thus have examined the change in NOx following treadmill exercise stress. In this study, 12 apparently healthy subjects showed an increase (from baseline) in venous NOx at peak effort and during recovery (12 ± 9.1 and 17 ± 15.3 μM respectively, P < 0.05). In contrast, 10 subjects with cardiovascular disease showed no significant increases. Additionally, a correlation between VO 2peak and the change in circulating NOx ( r 2 = 0.4585, P ≤ 0.01) indicated the subjects who could exercise hardest also produced the most NO.

Tailoring magnetic nanocrystal superlattices by chemical engineering

Recognizing the technological potential of magnetic nanocrystals for the next generation of high density data storage [S. Sun and C. B. Murray, J. Appl. Phys. 85, 4325 (1999)] and as markers in biological systems for detection and manipulation of attached molecules, [L. Lagae, R. Wirix-Speetjens, J. Das, D. Graham, H. Ferreira, P. P. F. Freitas, G. Borghs, and J. De Boeck, J. Appl. Phys. 91, 7445 (2002)] ligand stabilized nanoparticles prepared via high–temperature solution–phase synthesis [C. B. Murray, S. Sun, W. Gaschler, H. Doyle, T. A. Betley, and C. R. Kagan, IBM J. Res. Dev. 45, 47 (2001); V. F. Puntes, K. M. Krishnan, and A. P. Alivisatos, Science 291, 2115 (2001)] rapidly gain interest in research and development. Following a chemical preparation route recently proposed [V. F. Puntes, K. M. Krishnan, and A. P. Alivisatos, Science 291, 2115 (2001)] Co and FeCo alloyed nanocrystals have been synthesized with different nanocrystal size distributions employing different growth rates during preparation. The resulting magnetic properties are a function of the particle size distribution and can be tailored for superparamagnetic or ferromagnetic behavior at room temperature. In addition, the possibility of synthesizing FeCo alloyed nanoparticles is demonstrated and discussed.

Long-chain acyl-homoserine lactone quorum-sensing regulation of Rhodobacter capsulatus gene transfer agent production.

Many proteobacteria use acyl-homoserine lactones as quorum-sensing signals. Traditionally, biological detection systems have been used to identify bacteria that produce acyl-homoserine lactones, although the specificities of these detection systems can limit discovery. We used a sensitive approach that did not require a bioassay to detect production of long-acyl-chain homoserine lactone production by Rhodobacter capsulatus and Paracoccus denitrificans. These long-chain acyl-homoserine lactones are not readily detected by standard bioassays. The most abundant acyl-homoserine lactone was N-hexadecanoyl-homoserine lactone. The long-chain acyl-homoserine lactones were concentrated in cells but were also found in the culture fluid. An R. capsulatus gene responsible for long-chain acyl-homoserine lactone synthesis was identified. A mutation in this gene, which we named gtaI, resulted in decreased production of the R. capsulatus gene transfer agent, and gene transfer agent production was restored by exogenous addition of N-hexadecanoyl-homoserine lactone. Thus, long-chain acyl-homoserine lactones serve as quorum-sensing signals to enhance genetic exchange in R. capsulatus.

NANOSIZED OPTICAL BIOSENSORS

TINY SPECS OF SILVER COULD play a central role in a sensitive and inexpensive biological detection system, according to a study conducted at Northwestern University. The nanoparticle- based analytical procedure might be used to monitor ligand-receptor binding events, protein adsorption on self-assembled monolayers, and other types of analyte-surface interactions. Based on a technique in which researchers measure changes in the extinction (absorption plus scattering) spectrum of microscopic metal particles, the new procedure provides a way to measure low concentrations of specific analyte molecules using simple UV-Vis spectroscopy instrumentation. Chemistry professor Richard P. Van Duyne and graduate student Amanda J. Haes demonstrated the method by detecting low picomolar quantities of biotin-strep tavidin-a model ligand-receptor system [ J. Am. Chem. Soc. , published online Aug. 8, http://dx.doi.org/10.1021/ja020393x]. The detection principle is based on a variation of surface plasmon resonance (SPR) spe...

New applications advisable for gallium nitride

Abstract GaN-based visible light-emitting diodes and laser diodes are already commercialized for a variety of lighting and data storage applications. This materials system is also showing promise for microwave and high power electronics intended for radar, satellite, wireless base stations, and utility grid applications; for biological detection systems; and for a new class of spin-transport electronics (spintronics) in which the spin of charge carriers is exploited. The explosive increase of interest in the AlGaInN family of materials in recent years has been fueled by the application of blue/green/UV light-emitting diodes (LEDs) in full-color displays, traffic lights, automotive lighting, and general room lighting (using so-called white LEDs) 1 . In addition, blue/green laser diodes can be used in high storage-capacity digital versatile disk (DVD) systems 2 . AlGaN-based photodetectors are also useful for solar-blind UV detection and have applications as flame sensors for control of gas turbines or detection of missiles.

Bioterrorism preparedness: II: the community and emergency medical services systems

Disaster planning is an arduous task. Perhaps no form of disaster is more difficult to prepare for than one resulting from the intentional, covert release of a biological pathogen or toxin. The complexities of response operations and the perils of inadequate preparation cannot be overemphasized. Even with detailed planning, deviations from anticipated emergency operations plans are likely to occur. Several federal programs have been initiated to assist communities in enhancing their preparedness for events involving biological and other agents of mass destruction. Many of these, such as the Metropolitan Medical Response Systems (MMRS) Program [37,38], will be discussed elsewhere. Community preparedness will be enhanced by: 1. Implementing a real-time public health disease surveillance program linking local healthcare, emergency care, EMS, the CDC, local law enforcement, and the FBI 2. Improved real-time regional patient and healthcare capacity status management 3. Development of affordable, accurate biological agent detection systems 4. Incorporation of standardized education and training curricula (appropriate for audience) on terrorism and biological agents into healthcare training programs 5. Expansion of federal and state programs to assist communities in system development 6. Increased public awareness and education programs.

Testing a Biological Weapon Detection System in a Distributed Virtual Reality Environment

A study called Bio-Sim was sponsored by the U.S. Army to examine the LR-BSDS (Long-Range Biological Standoff Detection System), a laser-based biological cloud detector. Testing of the device was performed in a distributed virtual reality environment (DVRE). The three primary objectives of the study were to: 1) determine appropriate tactics, techniques, and procedures for use of the LR-BSDS, 2) gain a better understanding of those battlefield situations and characteristics that limit the effective use of the LR-BSDS, and 3) demonstrate the potential to use a DVRE simulation for training using the LR-BSDS. This paper will focus on the third of those objectives. The study consisted of three phases. The first phase involved calculating theoretical concentration limits of a dispersed biohazard as a function of time, dispersion concentration, and distance to sensor. The second phase resulted in a set of abbreviated bio-cloud detection missions run in a DVRE. The third phase was a set of full-length human-in-the-loop trial missions run by trained LR-BSDS operators and helicopter flight crews, using a DVRE and computer-based simulators for the LR-BSDS, helicopter, biocloud dispersion, and bio-cloud transport.

A new magnetic bead-based, filterless bio-separator with planar electromagnet surfaces for integrated bio-detection systems

A new filterless bio-separator separating magnetic microbeads from a carrier fluid has been designed, fabricated, and characterized as a core component of biological cell sampling and detecting systems. To maximize the sampling capability, a planar electromagnet surface with a serpentine coil and semi-encapsulated permalloy has been realized. Using this bio-separator, antibody-coated magnetic beads have been successfully separated from the bio-buffer suspension solution and characterized in dynamic fluid flow. The magnetic characteristics of the bio-separator have been simulated and experimentally studied to establish and validate the design rules for fabrication. The realized filterless bio-separator has a high potential in biomedical and biological detection systems. It is especially useful in selective sampling and separation of small amounts of bio-molecules (e.g., antigen) for on-chip micro total analysis systems (μ-TAS) or remote detection systems.