Validation of Microfluidic Hybridization Device for Post-PCR Analysis and Clinical Identification of Human Cytomegalovirus (CMV)
Abstract:This paper validates the microfluidics-based method of post-polymerase chain reaction (PCR) analysis by hybridization and simultaneous size separation of DNA. The usefulness of this technique is illustrated by the measurements of target 57 bp cytomegalovirus (CMV) amplicon that hybridized with the fluorescence-labeled peptide nucleic acid (PNA) probe used as the model analyte system. The hybrids formed were detected by laser-induced fluorescence detection system. A microfluidic device with serpentine microchannel pattern accommodated laminar flow and laminar secondary flow which enabled hybridization and separation of probe-bound DNA. Mixing by diffusion provided for optimum contact of the probe and target DNA to form double-stranded DNA (dsDNA). The dsDNAs were separated by diffusion according to their size, and secondary laminar flow at the channel curves. This approach expedited the hybridization reaction, increased hybridization efficiency, and achieved DNA complex separation by flow rate modulation and proper microchannel design. The validity of this microfluidic hybridization chip was evaluated by comparison with pp65 antigenemia assay. Results show the reliability of this microfluidic-based CMV detection as well as its potential as an alternative to antigenemia. We foresee that this method may also serve as a platform for the detection of other clinical pathogens.
Document Type: Research Article
Publication date: 2010-09-01
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