Toward a microchip-based solid-phase extraction method for isolation of nucleic acids

Abstract

A silica-based solid-phase extraction system suitable for incorporation into a microchip platform (nu-total analytical system; nu-TAS) would find utility in a variety of genetic analysis protocols, including DNA sequencing. The extraction procedure utilized is based on adsorption of the DNA onto bare silica. The procedure involves three steps: (i) DNA adsorption in the presence of a chaotropic salt, (ii) removal of contaminants with an alcohol/water solution, and (iii) elution of the adsorbed DNA in a small volume of buffer suitable for polymerase chain reaction (PCR) amplification. Multiple approaches for incorporation of this protocol into a microchip were examined with regard to extraction efficiency, reproducibility, stability, and the potential to provide PCR-amplifiable DNA. These included packing microchannels with silica beads only, generating a continuous silica network via sol-gel chemistry, and combinations of these. The optimal approach was found to involve immobilizing silica beads packed into the channel using a sol-gel network. This method allowed for successful extraction and elution of nanogram quantities of DNA in less than 25 min, with the DNA obtained in the elution buffer fraction. Evaluation of the eluted DNA indicated that it was of suitable quality for subsequent amplification by PCR.

Purification of Nucleic Acids in Microfluidic Devices

“…Wolfe et al extrapolated this technology to the microchip and evaluated a variety of silica and silica bead/sol-gel matrices for DNA purification. 14 However, immobilization of silica particles in the microchannels is problematic. These issues can be overcome with weir structures fabricated during the glass etching step with or without a sol-gel "glue" process to immobilize the silica beads.…”

Extraction of genomic DNA using a new amino silica monolithic column

“…Tian et al [9] packed silica beads in a capillary by first for DNA purification. As extension of this work, the silica beads were held in the microchip channel by weirs [10,11] or immobilized using sol -gel chemistry [12]. One potential problem was that the silica beads had the tendency to pack more tightly under flow, thus increasing the back pressure [10].…”

“…As extension of this work, the silica beads were held in the microchip channel by weirs [10,11] or immobilized using sol -gel chemistry [12]. One potential problem was that the silica beads had the tendency to pack more tightly under flow, thus increasing the back pressure [10]. The requirement of frits or weirs for the immobilization of particles also increased the difficulty in fabrica-tion.…”

On-Chip Integratable Elastomeric Dome Valves for Glass Microfluidic Systems

“…(13,14) In contrast, Borofloat glass-based microfluidic devices are better autofluorescent properties compare with plastic-based chips. (15) In addition, glassbased microfluidic devices have shown advantages in applications such as solid-phase extraction (SPE), (16) capillary electrophoresis (CE), (17) and high-sensitivity fluorescence detection by CE. (18) Since microfluidic devices (19) fabricated by glass-to-glass thermal bonding technology provide a permanent bond, glass-based microfluidic devices are applicable for specialized applications that require high pneumatic pressure and high temperature (e.g., polymerase chain reaction (PCR) thermocycling).…”