Detection of Respiratory Viruses with Plastic High Throughput Screening Devices

Abstract

ABSTRACTMicroarrays 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 respiratory pathogens: Influenza A virus, respiratory syncytial virus, human enterovirus, and human 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 point-of-care use that is based on a low-cost portable Micro-Total-Analysis-System (μ-TAS).