AVS 50th International Symposium
    Biomaterial Interfaces Thursday Sessions
       Session BI+SS-ThA

Paper BI+SS-ThA5
Pb@super 2+@ Sensitive Catalytic DNA Assay Integrated into Microfluidic Channels

Thursday, November 6, 2003, 3:20 pm, Room 318/319

Session: Biodiagnostics
Presenter: R.A. Zangmeister, National Institute of Standards and Technology
Authors: R.A. Zangmeister, National Institute of Standards and Technology
M. Tarlov, National Institute of Standards and Technology
Correspondent: Click to Email
Advances in microchip technology coupled with innovative bioassays are advancing the field of biosensors. We previously reported a method for immobilizing single-stranded DNA (ss-DNA) probe molecules in polyacrylamide hydrogels within plastic microfluidic channels. Spatially defined plugs are formed by photopolymerization of a solution containing 19:1 polyacrylamide/bisacrylamide and ss-DNA modified at the 5' end with an acrylic acid group. Low concentrations of fluorescent-tagged ss-DNA targets can be captured and detected in the hydrogels. We aim to couple this technology with a novel bioassay based on the response of catalytic DNA to Pb@super 2+@ ions in solution. It is reported to show > 80-fold selectivity for Pb@super 2+@ over other divalent metal ions, and with fluorescent tag modifiers can be used to detect Pb@super 2+@ ions over a large concentration range (10 nmol to 4 mmol).@footnote 1@ Our goal is to immobilize the enzyme strand sequence of the catalytic DNA duplex into the hydrogel plugs immobilized in microfluidic channels. Our strategy is to electrophorese fluorescently tagged substrate strands into the hydrogel plug where they hybridize with the immobilized enzyme strand to form the catalytic DNA system. Then Pb@super 2+@ is electrophoresed into the hydrogel plug resulting in the catalytic cleavage of the substrate strand and the release of the fluorescent-tagged sequence fragment that is detected using a fluorescence microscope. The combination of these two technologies results in a Pb@super 2+@ detection system with enhanced sensitivity due to the high loading of DNA probes in the hydrogel plug, the spatially confined, directed mass transfer characteristics of the microfluidic channels, and the inherently low fluorescent background of the hydrogels. The immobilization, retention of catalytic DNA activity, and current limits of detection will be discussed. @FootnoteText@@footnote 1@ Li, J.; Lu, Y., J. Am. Chem. Soc. 2000, 122, 10466-10467.