Toward an on-chip multiplexed nucleic acid hybridization assay using immobilized quantum dot-oligonucleotide conjugates and fluorescence resonance energy transfer

Semiconductor quantum dots (QD) are a class of NP with photophysical properties that are ideally suited for optical multiplexing and use as donors in fluorescence resonance energy transfer (FRET). A new strategy is presented for the development of multiplexed DNA hybridization assays using immobilized QDs in a microfluidic system. Green- or red-emitting QDs were immobilized via self-assembly with a multidentate-thiol-derivatized glass slide, and subsequently conjugated with amine-terminated probe oligonucleotides using carbodiimide activation. Immobilized QD-probe conjugates were then passivated with adsorbed non-complementary oligonucleotides to achieve selectivity in microfluidic assays. Target nucleic acid sequences hybridized with QD-probe conjugates and were labeled with Cy3 or Alexa Fluor 647 as acceptor dyes for the QD donors, where FRET-sensitized dye emission provided a signal for the detection of picomolar quantities of target. The simultaneous immobilization of green- and red-emitting QDs at different ratios within a microfluidic channel was demonstrated as a step toward multiplexed assays.

[1]  U. Krull,et al.  Multiplexed interfacial transduction of nucleic acid hybridization using a single color of immobilized quantum dot donor and two acceptors in fluorescence resonance energy transfer. , 2010, Analytical chemistry.

[2]  Igor L. Medintz,et al.  Self-assembled nanoscale biosensors based on quantum dot FRET donors , 2003, Nature materials.

[3]  W. Russ Algar,et al.  The application of quantum dots, gold nanoparticles and molecular switches to optical nucleic-acid diagnostics , 2009 .

[4]  Xiaogang Peng,et al.  Experimental Determination of the Extinction Coefficient of CdTe, CdSe, and CdS Nanocrystals , 2003 .

[5]  U. Krull,et al.  Multidentate surface ligand exchange for the immobilization of CdSe/ZnS quantum dots and surface quantum dot-oligonucleotide conjugates. , 2008, Langmuir : the ACS journal of surfaces and colloids.

[6]  U. Krull,et al.  Towards multi-colour strategies for the detection of oligonucleotide hybridization using quantum dots as energy donors in fluorescence resonance energy transfer (FRET). , 2007, Analytica chimica acta.

[7]  U. Krull,et al.  New opportunities in multiplexed optical bioanalyses using quantum dots and donor–acceptor interactions , 2010, Analytical and bioanalytical chemistry.

[8]  A. MacKenzie,et al.  Rapid detection of single nucleotide polymorphisms associated with spinal muscular atrophy by use of a reusable fibre-optic biosensor. , 2004, Nucleic acids research.

[9]  P. Chou,et al.  One-pot synthesis and characterization of high-quality CdSe/ZnX (X=S, Se) nanocrystals via the CdO precursor , 2004 .

[10]  S. Nie,et al.  Quantum dot bioconjugates for ultrasensitive nonisotopic detection. , 1998, Science.

[11]  U. Krull,et al.  Toward a multiplexed solid-phase nucleic acid hybridization assay using quantum dots as donors in fluorescence resonance energy transfer. , 2009, Analytical chemistry.

[12]  A. J. Tavares,et al.  Toward a solid-phase nucleic acid hybridization assay within microfluidic channels using immobilized quantum dots as donors in fluorescence resonance energy transfer , 2011, Analytical and bioanalytical chemistry.

[13]  U. Krull,et al.  Quantum dots as donors in fluorescence resonance energy transfer for the bioanalysis of nucleic acids, proteins, and other biological molecules , 2008, Analytical and bioanalytical chemistry.

[14]  Igor L. Medintz,et al.  Resonance Energy Transfer Between Luminescent Quantum Dots and Diverse Fluorescent Protein Acceptors. , 2009, The journal of physical chemistry. C, Nanomaterials and interfaces.

[15]  Xiaogang Peng,et al.  Formation of high-quality CdTe, CdSe, and CdS nanocrystals using CdO as precursor. , 2001, Journal of the American Chemical Society.

[16]  Ulrich J Krull,et al.  Developing mixed films of immobilized oligonucleotides and quantum dots for the multiplexed detection of nucleic acid hybridization using a combination of fluorescence resonance energy transfer and direct excitation of fluorescence. , 2010, Langmuir : the ACS journal of surfaces and colloids.

[17]  Bernhardt L Trout,et al.  Modified ligand-exchange for efficient solubilization of CdSe/ZnS quantum dots in water: a procedure guided by computational studies. , 2008, Langmuir : the ACS journal of surfaces and colloids.

[18]  U. Krull,et al.  Adsorption and hybridization of oligonucleotides on mercaptoacetic acid-capped CdSe/ZnS quantum dots and quantum dot-oligonucleotide conjugates. , 2006, Langmuir : the ACS journal of surfaces and colloids.

[19]  Xuezhu Liu,et al.  Electrokinetically based approach for single-nucleotide polymorphism discrimination using a microfluidic device. , 2005, Analytical chemistry.

[20]  Ulrich J Krull,et al.  Beyond labels: a review of the application of quantum dots as integrated components of assays, bioprobes, and biosensors utilizing optical transduction. , 2010, Analytica chimica acta.

[21]  Xuezhu Liu,et al.  Electrokinetically controlled DNA hybridization microfluidic chip enabling rapid target analysis. , 2004, Analytical chemistry.

[22]  W. Russ Algar,et al.  Toward a hybridization assay using fluorescence resonance energy transfer and quantum dots immobilized in microfluidic channels , 2010, Photonics North.

[23]  Igor L. Medintz,et al.  Quantum dot bioconjugates for imaging, labelling and sensing , 2005, Nature materials.

[24]  U. Krull,et al.  Interfacial transduction of nucleic acid hybridization using immobilized quantum dots as donors in fluorescence resonance energy transfer. , 2009, Langmuir : the ACS journal of surfaces and colloids.