Molecular Beacon Lighting up on Graphene Oxide

Abstract

A molecular beacon (MB) is comprised of a fluorophore and a quencher linked by a DNA hairpin. MBs have been widely used for homogeneous DNA detection. In addition to molecular quenchers, many nanomaterials such as graphene oxide (GO) also possess excellent quenching efficiency. Most reported fluorescent sensors relied on DNA probes physisorbed by GO, which may suffer from non-specific probe displacement and false positive signal. In this work, we report the preparation and characterization of a MB using graphene oxide (GO) as quencher, where an amino and FAM (6carboxyfluorescein) dual labeled DNA was covalently attached to GO via an amide linkage. A major challenge was to remove noncovalently attached probes due to strong DNA adsorption by GO. While DNA desorption was favored at low salt, high pH, high temperature, and by using organic solvents, the complementary DNA was required to achieve complete desorption of non-covalently linked DNA probes. The DNA adsorption energy was measured using isothermal titration calorimetry, revealing the heterogeneous nature of GO. The covalent probe has a detection limit of 2.2 nM using a sample volume of 0.05 mL. With 2 mL sample, the detection limit can reach 150 pM. The covalent probe is highly resistant to non-specific probe displacement and will find applications in serum and cellular samples where high probe stability is demanded.

Effects of Polyethylene Glycol on DNA Adsorption and Hybridization on Gold Nanoparticles and Graphene Oxide

“…The GO-DNA complex was washed extensively to remove non-covalently attached DNA as detailed in a previous publication. 47 Finally the complex was dispersed in buffer (25 mM HEPES buffer, pH 7.6, 150 mM NaCl, 1 mM MgCl2) and stored at 4 °C with a final GO concentration of 100 μg/mL. DNA hybridization to AuNPs and GO.…”

Comparison of Graphene Oxide and Reduced Graphene Oxide for DNA Adsorption and Sensing

“…[6][7][8][9][10][11] In addition, amino-modified DNAs were covalently attached to the carboxyl groups on GO forming an amide bond, avoiding non-specific probe displacement. [12][13][14][15] Many DNA-related enzymes were also involved to introduce functions such as signal amplification. 16,17 Finally, DNA/GO conjugates were used to template materials synthesis such as metal nanoparticles, [18][19][20] and stacked GO sheets.…”

Fast and Single-step Fluorescence-based Competitive Bioassay Microdevice Combined PDMS Microchannel Arrays Separately Immobilizing Graphene Oxide-Analyte Conjugates and Fluorescently-labelled Receptor Proteins

“…Graphene oxide (GO), which is a two-dimensional carbon nanomaterial, has recently gained attention because of its exceptionally high quenching ability based on fluorescence resonance energy transfer (FRET), 17 and this property has been applied in DNA sensors, 18,19 immunosensors, [20][21][22] and aptasensors. [23][24][25][26][27] Furthermore, recently, we reported a capillarytype homogeneous immunoassay microdevice based on the fluorescence quenching function of the GO immobilizing antibody 13 and a heterogeneous immunoassay microdevice based on the size separation and fluorescence quenching functions of GO-containing hydrogel.…”