Chemical Sensors Group, Department of Chemical and Physical Sciences, University of Toronto Mississauga, 3359 Mississauga Road North, Mississauga, Ontario L5L 1C6, Canada.
Anal Chem. 2012 Jan 3;84(1):312-9. doi: 10.1021/ac2025943. Epub 2011 Dec 19.
The glass surface of a glass-polydimethylsiloxane (PDMS) microfluidic channel was modified to develop a solid-phase assay for quantitative determination of nucleic acids. Electroosmotic flow (EOF) within channels was used to deliver and immobilize semiconductor quantum dots (QDs), and electrophoresis was used to decorate the QDs with oligonucleotide probe sequences. These processes took only minutes to complete. The QDs served as energy donors in fluorescence resonance energy transfer (FRET) for transduction of nucleic acid hybridization. Electrokinetic injection of fluorescent dye (Cy3) labeled oligonucleotide target into a microfluidic channel and subsequent hybridization (within minutes) provided the proximity for FRET, with emission from Cy3 being the analytical signal. The quantification of target concentration was achieved by measurement of the spatial length of coverage by target along a channel. Detection of femtomole quantities of target was possible with a dynamic range spanning an order of magnitude. The assay provided excellent resistance to nonspecific interactions of DNA. Further selectivity of the assay was achieved using 20% formamide, which allowed discrimination between a fully complementary target and a 3 base pair mismatch target at a contrast ratio of 4:1.
玻璃-聚二甲基硅氧烷（PDMS）微流控通道的玻璃表面经过修饰，开发出一种用于定量测定核酸的固相分析。通道内的电渗流（EOF）用于输送和固定半导体量子点（QD），电泳用于将寡核苷酸探针序列修饰到 QD 上。这些过程仅需几分钟即可完成。QD 作为荧光共振能量转移（FRET）中的能量供体，用于核酸杂交的转导。将荧光染料（Cy3）标记的寡核苷酸靶标电动力学注入微流控通道中，随后进行杂交（几分钟内），提供了 FRET 的接近程度，Cy3 的发射作为分析信号。通过测量目标沿通道的覆盖长度来实现目标浓度的定量。可以检测到飞摩尔数量级的目标，动态范围跨越一个数量级。该分析提供了对 DNA 非特异性相互作用的出色抗性。通过使用 20%甲酰胺进一步提高了分析的选择性，这允许在对比度为 4:1 的情况下区分完全互补的靶标和 3 个碱基对错配的靶标。
