A simple, valveless microfluidic sample preparation device for extraction and amplification of DNA from nanoliter-volume samples.
A glass microdevice has been constructed for the on-line integration of solid-phase extraction (SPE) of DNA and polymerase chain reaction (PCR) on a single chip. The chromatography required for SPE in the microfluidic sample preparation device (muSPD) was carried out in a silica bead/sol-gel SPE bed, where the purified DNA was eluted directly into a downstream chamber where conventional thermocycling allowed for PCR amplification of specific DNA target sequences. Through rapid, simple passivation of the PCR chamber with a silanizing reagent, reproducible DNA extraction and amplification was demonstrated from complex biological matrixes in a manner amenable to any research laboratory, using only a syringe pump and a conventional thermocycler. The muSPD allowed for SPE concentration of DNA from 600 nL of blood coupled to subsequent on-chip amplification that yielded a detectable amplicon; this simple device can be applied to a variety of routine genetic analyses without the need for sophisticated instrumentation. In addition, the applicability of these developments to nonconventional thermocycling was demonstrated through the use of noncontact, IR-mediated heating. This was exemplified with the isolation of DNA from an anthrax spore-spiked nasal swab and the subsequent on-chip amplification of target DNA sequences in a total processing time of only 25 min.
Legendre
Lindsay A
LA
Department of Chemistry, University of Virginia, Charlottesville, Virginia 22904, USA.
Bienvenue
Joan M
JM
Roper
Michael G
MG
Ferrance
Jerome P
JP
Landers
James P
JP
D004247:DNA / Q000302:isolation & purification*; D046210:Microfluidic Analytical Techniques / Q000295:instrumentation* / Q000379:methods; D009862:Online Systems; D016133:Polymerase Chain Reaction / Q000295:instrumentation* / Q000379:methods
D004247:DNA