A microbead-incorporated centrifugal sample pretreatment microdevice.

Abstract

In this study, we present a novel bead-incorporated centrifugal sample pretreatment microdevice to purify influenza A H3N2 viral RNA. Simple revolution per minute (RPM) control can lead to RNA capture on a bead-bed, and the sequential loading of a washing solution and an elution solution. Tetraethoxy orthosilicate (TEOS)-treated glass microbeads were utilized as a capture matrix. The sample pretreatment microdevice consists of four reservoirs for storing an RNA sample, a washing solution, an elution solution, and a collected sample, and they were merged at the microbead-bed microchannel. The washing solution reservoir and the elution solution reservoir were connected to the bead-bed microchannel through a capillary valve and a siphon channel, respectively. An RNA sample (a lysed influenza A H3N2 virus), a washing solution (70% ethanol) and an elution solution (water or a reverse transcription-polymerase chain reaction (RT-PCR) cocktail) were loaded into the designated reservoirs, and they were successively transported to the bead-bed by RPM control owing to the optimized channel design. Purified RNAs could be obtained in 440 s. Then, a target H3 gene was amplified by an off-chip based real-time RT-PCR to evaluate the capture efficiency of RNA on our proposed microdevice. 81% of RNAs were successfully captured and purified. Interestingly, the use of the RT-PCR cocktail itself as an elution solution resulted in a 76% capture yield. Furthermore, we successfully performed RNA purification from the clinical nasopharyngeal swabs to identify the subtype of the influenza A virus. This platform provides high potential for the direct integration of the sample pretreatment microdevice into the downstream micro-PCR unit to create a total genetic analysis microsystem.

• Authors: Jae Hwan Jung, Byung Hyun Park, Young Ki Choi, Tae Seok Seo
• Journal: Lab on a chip (Lab Chip) Vol. 13 Issue 17 Pg. 3383-8 (Sep 07 2013) ISSN: 1473-0189 [Electronic] England
• PMID: 23824467 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)

Chemical References

• RNA, Viral

Topics

• Analytic Sample Preparation Methods (instrumentation)
• Centrifugation (instrumentation)
• Humans
• Influenza A Virus, H3N2 Subtype
• Microfluidic Analytical Techniques (instrumentation)
• Microspheres
• Polymerase Chain Reaction
• RNA, Viral (chemistry, genetics, isolation & purification)