title: Enhancing the speed of morpholino-DNA biosensor by electrokinetic concentration of DNA in a microfluidic chip.
abstract: Electrokinetic methods that conveniently concentrate charged analytes by orders of magnitude are highly attractive for nucleic acid assays where they can bypass the complexity and costs of enzyme-based amplification. The present study demonstrates an electrokinetic concentration device incorporating charge-neutral morpholino (MO) probes: as DNA analyte is concentrated in a microfluidic channel using ion concentration polarization (ICP) it is simultaneously hybridized to spots of complementary MO probes immobilized on the channel floor. This approach is uniquely favored by the match between the optimum buffer ionic strength of approximately 10mM for both MO-DNA surface hybridization and electrokinetic concentration. The simple and easily scalable poly(dimethylsiloxane) (PDMS) microfluidic device was fabricated using soft lithography and contact printing of a conductive polymer, poly(3,4-ethylenedioxythiophene)-polystyrene sulfonate (
  PSS) as a cation-selective membrane material. Using the microfluidic concentrator, we could increase the concentration of DNA by three orders of magnitude in less than 5 min at an electric field of 75 Vcm(-1). The 1000-fold increase in concentration of DNA led to an increase in the speed of MO-DNA hybridization by two orders of magnitude and enabled a detection sensitivity of ~1 nM within 15 min of concentration. Using the proposed microfluidic concentrator, we also demonstrated a rapid hybridization with a binary DNA mixture, containing a fully complementary and a non-complementary sequence to mimic molecular backgrounds present in real DNA samples.
authors.affiliation: Division of Engineering, New York University Abu Dhabi, P.O. Box 129188, Abu Dhabi, UAE.
authors.forename: Diogo
authors.initials: D
authors.lastname: Martins
authors.affiliation: Department of Chemical and Biomolecular Engineering, New York University Polytechnic School of Engineering, Brooklyn, NY 11201, USA.
authors.forename: Rastislav
authors.initials: R
authors.lastname: Levicky
authors.affiliation: Division of Engineering, New York University Abu Dhabi, P.O. Box 129188, Abu Dhabi, UAE; Department of Chemical and Biomolecular Engineering, New York University Polytechnic School of Engineering, Brooklyn, NY 11201, USA. Electronic address: rafael.song@nyu.edu.
authors.forename: Yong-Ak
authors.initials: YA
authors.lastname: Song
mesh_terms: D001483:Base Sequence; D015374:Biosensing Techniques / Q000295:instrumentation*; D004247:DNA / Q000032:analysis* / Q000302:isolation & purification; D004560:Electricity; D004867:Equipment Design; D007700:Kinetics; D057230:Limit of Detection; D046210:Microfluidic Analytical Techniques / Q000295:instrumentation*; D060172:Morpholinos / Q000737:chemistry*; D009693:Nucleic Acid Hybridization; D020411:Oligonucleotide Array Sequence Analysis / Q000295:instrumentation; D011137:Polystyrenes / Q000737:chemistry*; D013876:Thiophenes / Q000737:chemistry*
chemical_list: D060172:Morpholinos; D011137:Polystyrenes; D013876:Thiophenes; C533756:poly(3,4-ethylenedioxythiophene)-poly(styrenesulfonate); D004247:DNA
keywords: DNA; Hybridization; Ion polarization concentration (ICP); Microfluidics; Morpholino; Poly(3,4-ethylenedioxythiophene)-polystyrene sulfonate (PEDOT:PSS)