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Benjamin J Hindson,
Dora M Gutierrez,
Kevin D Ness,
Anthony J Makarewicz, Thomas R Metz,
Ujwal S Setlur,
William B Benett,
Jeffrey M Loge,
Bill W Colston,
Paul S Francis,
Neil W Barnett,
John M Dzenitis
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ABSTRACT: We present a fully automated DNA purification module comprised of a micro-fabricated chip and sequential injection analysis system that is designed for use within autonomous instruments that continuously monitor the environment for the presence of biological threat agents. The chip has an elliptical flow channel containing a bed (3.5 x 3.5 mm) of silica-coated pillars with height, width and center-to-center spacing of 200, 15, and 30 microm, respectively, which provides a relatively large surface area (ca. 3 cm(2)) for DNA capture in the presence of chaotropic agents. We have characterized the effect of various fluidic parameters on extraction performance, including sample input volume, capture flow rate, and elution volume. The flow-through design made the pillar chip completely reusable; carryover was eliminated by flushing lines with sodium hypochlorite and deionized water between assays. A mass balance was conducted to determine the fate of input DNA not recovered in the eluent. The device was capable of purifying and recovering Bacillus anthracis genomic DNA (input masses from 0.32 to 320 pg) from spiked environmental aerosol samples, for subsequent analysis using polymerase chain reaction-based assays.
The Analyst 03/2008; 133(2):248-55. · 4.23 Impact Factor
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Benjamin J Hindson,
Mary T McBride,
Anthony J Makarewicz,
Bruce D Henderer,
Ujwal S Setlur,
Sally M Smith,
Dora M Gutierrez, Thomas R Metz,
Shanavaz L Nasarabadi,
Kodumudi S Venkateswaran,
Stephen W Farrow,
Bill W Colston,
John M Dzenitis
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ABSTRACT: The autonomous pathogen detection system (APDS) is an automated, podium-sized instrument that continuously monitors the air for biological threat agents (bacteria, viruses, and toxins). The system has been developed to warn of a biological attack in critical or high-traffic facilities and at special events. The APDS performs continuous aerosol collection, sample preparation, and detection using multiplexed immunoassay followed by confirmatory PCR using real-time TaqMan assays. We have integrated completely reusable flow-through devices that perform DNA extraction and PCR amplification. The fully integrated system was challenged with aerosolized Bacillus anthracis, Yersinia pestis, Bacillus globigii, and botulinum toxoid. By coupling highly selective antibody- and DNA-based assays, the probability of an APDS reporting a false positive is extremely low.
Analytical Chemistry 02/2005; 77(1):284-9. · 5.86 Impact Factor
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Benjamin J Hindson,
Steve B Brown,
Graham D Marshall,
Mary T McBride,
Anthony J Makarewicz,
Dora M Gutierrez,
Duane K Wolcott, Thomas R Metz,
Ramakrishna S Madabhushi,
John M Dzenitis,
Billy W Colston
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ABSTRACT: An automated sample preparation module, based upon sequential injection analysis (SIA), has been developed for use within an autonomous pathogen detection system. The SIA system interfaced aerosol sampling with multiplexed microsphere immunoassay-flow cytometric detection. Metering and sequestering of microspheres using SIA was found to be reproducible and reliable, over 24-h periods of autonomous operation. Four inbuilt immunoassay controls showed excellent immunoassay and system stability over five days of unattended continuous operation. Titration curves for two biological warfare agents, Bacillus anthracis and Yersinia pestis, obtained using the automated SIA procedure were shown to be similar to those generated using a manual microtiter plate procedure.
Analytical Chemistry 08/2004; 76(13):3492-7. · 5.86 Impact Factor
