Rapid - PCR ( LightCycler ) in diagnosis of biological agents

Taleski, Vaso and Hadfield, Ted (2001) Rapid - PCR ( LightCycler ) in diagnosis of biological agents. In: The First World Congress on Chemical and Biological Terrorism, 21-27 April 2001, Dubrovnik, Croatia.

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Abstract

Although the historic use of biological weapons has been infrequent, a belief that state sponsored armies or terrorist organizations will use this type of weapon has never been greater which demands a capability for rapid medical response and early intervention. Molecular diagnostic methods, based on DNA amplification known as PCR (Polymerase Chain Reaction) are promising tools in fast and specific detection and identification of biological agent(s). The R.A.P.I.D.TM - PCR ( Ruggedized Advanced Pathogen Identification Device ) (fig.1), is a 32 sample capacity, automated instrument integrating Idaho Technology's LightCycler® technology into a portable, impact resistant package. This allows field identification of pathogens quickly. Monitoring the fluorescence from the double-stranded DNA dye SYBR® Green (fig.2), followed by differentiation of products by melting curves or from TaqMan® probes (6-FAM-oligo-TAMRA,), allows inexpensive quantification of low initial template copy number. Cycle sequencing reactions done in the thermocycling module of the R.A.P.I.D. system are faster, cleaner and more readable than parallel reactions done in a conventional heat block cycler. The use of air as the cycling medium ensures temperature uniformity and rapid heat exchange with the sample loaded in thin micro-capillary tubes which are ideally suited to temperature cycling, because of their extremely high surface area to volume ratio. A conventional PCR protocol takes up-to 3 hours to do 30 three temperature cycles. The R.A.P.I.D. system can complete a 40 cycle reaction in less than 20 minutes ( 6 to 30 min.). This makes R.A.P.I.D. system the fastest thermal cycler in the world. The United States Air Force has developed over 50 assays for infectious agents on the R.A.P.I.D. system. Assays for infectious agents typically consist of two temperature cycling for 40 cycles. Protocols for isolation of bacteria and viral DNA (or RNA) have been developed for clinical specimens, air samples and water samples. Protocols for food samples are being developed now. Assays in use for : Bacillus anthracis, Yersinia pestis, botulinum toxin, staphylococcal enterotoxins, Francisella tularensis, Salmonella, Shigella, Vibrio cholerae, E. coli, Campylobacter, VEE, West Nile, Yellow Fever, Brucella spp., and many others. The continuing effort and advances in testing design make this capability an asset for Army Commanders and Medical facilities alike.

Item Type: Conference or Workshop Item (Lecture)
Subjects: Medical and Health Sciences > Health sciences
Medical and Health Sciences > Other medical sciences
Divisions: Faculty of Medical Science
Depositing User: Vaso Taleski
Date Deposited: 06 Sep 2013 09:54
Last Modified: 06 Sep 2013 09:54
URI: http://eprints.ugd.edu.mk/id/eprint/5444

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