Detection of Respiratory Viruses with Plastic High Throughput Screening Devices
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Detection of Respiratory Viruses with Plastic High Throughput Screening Devices Zhengshan Zhao1, Gerardo A. Diaz-Quijada2, Régis Peytavi1, Éric LeBlanc1, Johanne Frenette1, Guy Boivin1, Jim V. Zoval3, Marc Madou3, Michel M. Dumoulin2, Teodor Veres2, and Michel G. Bergeron1 1 Centre de recherche en infectiologie, Université Laval, 2705 Boul. Laurier, Sainte Foy, G1V 4G2, Canada 2 Industrial Materials Institute, National Research Council of Canada, 75 de Mortagne, Boucherville, J4B 6Y4, Canada 3 Department of Mechanical and Aerospace Engineering, University of California, Irvine, CA, 92697-3975
ABSTRACT Microarrays have become one of the most convenient tools for high throughput screening and have catalyzed major advances in genomics and proteomics. Other important applications can be found in medical diagnostics, detection of biothreats, drug discovery, etc. Integration of microarrays with microfluidic devices can be highly advantageous in terms of portability, shorter analysis time and lower consumption of expensive biological analytes. Since fabrication of microfluidic devices using traditional materials such as glass is rather expensive, there is a high interest in employing polymeric materials as a low cost alternative suitable for mass production. We present proof-of-concept DNA arrays on a plastic platform for the detection of four important human respiratory pathogens: influenza A virus, respiratory syncytial virus, enterovirus, and metapneumovirus. This was accomplished by amplifying the genetic material from the viruses and simultaneously labeling the amplicons with a fluorescent dye (Cy3) via a highly sensitive multiplex Reverse Transcription Polymerase Chain Reaction (RT-PCR). The resultant RT-PCR product was hybridized, without further purification, with an array of specific oligonucleotide probes (20 mers) that had been covalently bound to a plastic substrate. Results indicate a high signal to background ratio that is comparable to commercially available microarray glass slides. In addition, 5 minute hybridization on this plastic substrate has been demonstrated using a centrifugal microfluidic platform, paving the way to a rapid medical diagnostic device for pointof-care use that is based on a low-cost portable Micro-Total-Analysis-System (µ-TAS). INTRODUCTION Acute viral upper respiratory diseases are considered to be the most common illness for children in developed countries. Furthermore, these diseases represent a leading cause of mortality in developing countries around the world. Examples of common viruses responsible for respiratory infections are Influenza A (Flu A), respiratory syncytial virus (RSV), human metapneumo viruses (HMPV), and human enteroviruses (HEV). Due to the various etiologies of the upper respiratory tract infections, it is difficult to establish precisely the nature of the infection without performing a diagnostic method such as cell cultures. As a result of the length of time required
for cultures, antibiotic therapy is often prescribed in an empirical manner w
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