A disposable laser print-cut-laminate polyester microchip for multiplexed PCR via infra-red-mediated thermal control

Autor(es)

Ouyang, Yiwen ^[1] ; Duarte, Gabriela R. M. ^{[1, 2]} ; Poe, Brian L. ^[1] ; Riehl, Paul S. ^[1] ; dos Santos, Fernando M. ^[3] ; Martin-Didonet, Claudia C. G. ^[3] ; Carrilho, Emanuel ^{[4, 5]} ; Landers, James P. ^{[1, 6, 7]}

Afiliação do(s) autor(es)

^[1] Univ Virginia, Dept Chem, Charlottesville, VA 22904 - USA

^[2] Univ Fed Goias, BR-74690900 Goiania, Go - Brazil

^[3] Univ Estadual Goias, BR-75132400 Anapolis, Go - Brazil

^[4] Univ Sao Paulo, Inst Quim Sao Carlos, BR-13566590 Sao Carlos, SP - Brazil

^[5] Inst Nacl Ciencia & Tecnol Bioanalit, BR-13083970 Campinas, SP - Brazil

^[6] Univ Virginia, Dept Mech Engn, Charlottesville, VA 22904 - USA

^[7] Univ Virginia, Hlth Sci Ctr, Dept Pathol, Charlottesville, VA 22904 - USA

• Tipo de documento: Artigo Científico
• Fonte: Analytica Chimica Acta; v. 901, p. 59-67, DEC 11 2015.
• Citações Web of Science: 12

Resumo

Infrared (IR)-mediated thermal cycling system, a method proven to be a effective for sub-mu L scale polymerase chain reaction (PCR) on microchips, has been integrated with DNA extraction and separation on a glass microchip in a fully integrated micro Total Analysis System by Easley et al., in 2006. IR-PCR has been demonstrated on both glass and PMMA microdevices where the fabrication (bonding) is not trivial. Polyester-toner (PeT) microfluidic devices have significant potential as cost-effective, disposable microdevices as a result of the ease of fabrication (similar to\$0.25 USD and <10 min per device) and availability of commercial substrates. For the first time, we demonstrate here the thermal cycling in PeT microchips on the IR-PCR system. Undesirable IR absorption by the black-toner bonding layer was eliminated with a spatial filter in the form of an aluminum foil mask. The solution heating rate for a black PeT microchip using a tungsten lamp was 10.1 +/- 0.7 degrees C s(-1) with a cooling rate of roughly -12 +/- 0.9 degrees C s(-1) assisted by forced air cooling. Dynamic surface passivation strategies allowed the successful amplification of a 520 bp fragment of the lambda-phage genome (in 11 min) and a 1500 bp region of Azospirillum brasilense. Using a centrosymmetric chamber configuration in a multichamber PeT microchip, homogenous temperature distribution over all chambers was achieved with inter-chamber temperature differences at annealing, extension and denaturing steps of less than +/- 2 degrees C. The effectiveness of the multichamber system was demonstrated with the simultaneous amplification of a 390 bp amplicon of human beta-globin gene in five PeT PCR microchambers. The relative PCR amplification efficiency with a human beta-globin DNA fragment ranged from 70% to 90%, in comparison to conventional thermal cyclers, with an inter-chamber standard deviation of similar to 10%. Development of PeT microchips for IR-PCR has the potential to provide rapid, low-volume amplification while also integrating PCR with extraction upstream and separation/detection downstream. (C) 2015 Published by Elsevier B.V. (AU)

• Processo FAPESP: 05/04473-4 - Análises genéticas em sistemas nanofabricados
• Beneficiário: Gabriela Rodrigues Mendes Duarte
• Modalidade de apoio: Bolsas no Brasil - Doutorado