Convectively driven polymerase chain reaction thermal cycler

We have fabricated a low-cost disposable polymerase chain reaction thermal chamber that uses buoyancy forces to move the sample solution between the different temperatures necessary for amplification. Three-dimensional, unsteady finite element modeling and a simpler 1-D steady-state model are used together with digital particle image velocimetry data to characterize the flow within the device. Biological samples have been amplified using this novel thermal chamber. Time for amplification is less than 30 min. More importantly, an analysis of the energy consumption shows significant improvements over current technology.     

Status and cockpit dynamics: A review and empirical study.

One of the most troublesome dynamics evident in the airplane cockpit is related to patterns of authority relations between the captain and the first officer. Too often, captains fail to listen and first officers fail to speak. The authors propose that many instances of superordinate and subordinate behavior in the cockpit—the captain's tendency to reject input from other team members and the first officer's hesitancy to question the captain—represent cases of status generalization. First, the authors describe the theory of status generalization and show support for the operation of the theory by presenting examples of flightcrew behavior that the theory predicts. Second, an initial empirical test was conducted to instantiate the claim that captain–first officer differences can be seen as status differences. Finally, the significance and implications of this perspective are discussed. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Raman spectroscopy of Kevlar 49 fibre

The Raman spectrum of poly(p-phenylene terephthalate) fibre, Kevlar 49, is reported. Fibre which had been exposed to heat, stress and radiation indicated that within experimental error, no general change takes place in the chemical structure of the material as a whole. However the spectrum of a filament under stress shows that all bands in the spectrum become increasingly polarized with increased stress. The spectral characteristics of the stress fibre lead to a possible explanation of the polymer's response to deformation — opening of the angles in the amide linkage of the polymeric chain and improved alignment of crystallites along the fibre axis.     

Optical techniques for identifying animal and plant pathogens

Abstract

The contemporary revolution in biochemistry, bio-physics, and medicine is leading to an understanding of how biological entities operate in varied environments. One consequence is the increased understanding of antibodies, fluorescence markers, special substrates, and uses of the polymerase chain reaction process with biological markers. In particular, these efforts are leading to pathogen detection systems that are lower in cost, more biochemically specific, more accurate, faster, smaller, less demanding of infrastructure, and more accessible to more people than ever before. Most of these systems use optical techniques. Examples of miniature, portable detection systems based upon nucleotide amplification and on fluorescence sorting, concentration, and detection are discussed. These are applicable to the detection of pathogens and related toxins that appear in medical, agricultural, and national security situations confronting us today.     

Remote Detection of Organochlorides With a Fiber Optic Based Sensor II. a Dedicated Portable Fluorimeter

ABSTRACT

A dedicated portable fluorimeter, for use with fiber optic chemical sensors (FOCS) has been designed, constructed, tested, and calibrated. This represents a major advance in the development of a FOCS system suitable for in-field use. The portable fluorimeter uses an incandescent lamp, instead of a laser, for FOCS excitation and a photodiode, in place of a photomultiplier tube for detecting the fluorescence signal. It uses an optical system which is internally connected to a unitized optical block by 600 μm core optical fibers, to minimize alignment problems and increase overall system ruggedness. The system noise is less than .1.5 mV and the long-term drift is less than ±2 mV/hour. Measurements of organochloride were made at concentrations as low as 80 parts-per-billion with a signal to noise ratio of 40:1.     

Field-deployable sniffer for 2,4-dinitrotoluene detection

A field-deployable instrument has been developed to detect low-level 2,4-dinitrotoluene (2,4-DNT) vapors. The system is based on previously developed artificial nose technology and employs an array of sensory materials attached to the distal tips of an optical fiber bundle. Both semiselective and nonspecific, cross-reactive sensors were employed. Each sensor within the array responds differentially to vapor exposure so the array's fluorescence response patterns are unique for each analyte. The instrument is computationally "trained" to discriminate target response patterns from nontarget and background environments. This detection system has been applied to detect 2,4-DNT, an analyte commonly detected on the soil surface above buried 2,4,6-trinitrotoluene (TNT) land mines, in spiked soil and aqueous and ground samples. The system has been characterized and demonstrated the ability to detect 120 ppb 2,4-DNT vapor in blind (unknown) humidified samples during a supervised field test.     

A minisonicator to rapidly disrupt bacterial spores for DNA analysis.

Concerns about the use of anthrax spores as a weapon of mass destruction have motivated the development of portable instruments capable of detecting and monitoring a suspected release of the agent. Optimal detection of bacterial spores by PCR requires that the spores be disrupted to make the endogenous DNA available for amplification. The entire process of spore lysis, PCR, and detection can take several hours using conventional methods and instruments. In this report, a minisonicator and prototype spore lysis cartridge were built to disrupt Bacillus spores in 30 s for rapid, real-time PCR analysis. Utilization of the minisonicator improved PCR analysis by decreasing the limit of detection, reducing the time of detection, and increasing the signal amplitude. Total time of spore disruption and detection using the minisonicator and a microchip PCR instrument was less than 15 min.     

Autonomous detection of aerosolized Bacillus anthracis and Yersinia pestis

We have developed and tested a fully autonomous pathogen detection system (APDS) capable of continuously monitoring the environment for airborne biological threat agents. The system is designed to provide early warning to civilians in the event of a terrorist attack. The final APDS will be completely automated, offering aerosol sampling, in-line sample preparation fluidics, multiplexed detection and identification immunoassays, and orthogonal, multiplexed PCR (nucleic acid) amplification and detection. The system performance (current capabilities include aerosol collection, multiplexed immunoassays, sample archiving, data reporting, and alarming) was evaluated in a field test conducted in a Biosafety Level 3 facility, where the system was challenged with, and detected, a series of aerosolized releases containing two live, virulent biological threat agents (Bacillus anthracis and Yersinia pestis). Results presented here represent the first autonomous, simultaneous measurement of these agents.     

A reusable flow-through polymerase chain reaction instrument for the continuous monitoring of infectious biological agents

Continuous monitoring of the environment for infectious diseases and related biowarfare agents requires the implementation of practical cost-effective methodologies that are highly sensitive and specific. One compatible method employed in clinical diagnostics is real-time polymerase chain reaction (PCR) analysis. The utility of this technique for environmental monitoring is limited, however, by the utilization of single-use consumables in commercial PCR instruments. This greatly increases mechanical complexity, because sophisticated robotic mechanisms must replenish the disposable elements. An alternative strategy develops an autonomous monitoring system consisting of reusable modules that readily interface with fluidic circuitry in a flow-through scheme. The reduced complexity should increase reliability while decreasing operating costs. In this report, we describe a reusable, flow-through PCR module that functions as one component in such a system. This module was rigorously evaluated with Bacillus anthracis genomic DNA and demonstrated high repeatability, sensitivity, and efficiency, with no evidence of sample-to-sample carryover.     

High-throughput droplet digital PCR system for absolute quantitation of DNA copy number

Digital PCR enables the absolute quantitation of nucleic acids in a sample. The lack of scalable and practical technologies for digital PCR implementation has hampered the widespread adoption of this inherently powerful technique. Here we describe a high-throughput droplet digital PCR (ddPCR) system that enables processing of ~2 million PCR reactions using conventional TaqMan assays with a 96-well plate workflow. Three applications demonstrate that the massive partitioning afforded by our ddPCR system provides orders of magnitude more precision and sensitivity than real-time PCR. First, we show the accurate measurement of germline copy number variation. Second, for rare alleles, we show sensitive detection of mutant DNA in a 100,000-fold excess of wildtype background. Third, we demonstrate absolute quantitation of circulating fetal and maternal DNA from cell-free plasma. We anticipate this ddPCR system will allow researchers to explore complex genetic landscapes, discover and validate new disease associations, and define a new era of molecular diagnostics.