Vivek V. Thacker

DNA origami based assembly of gold nanoparticle dimers for surface-enhanced Raman scattering

Plasmonic sensors are extremely promising candidates for label-free single-molecule analysis but require exquisite control over the physical arrangement of metallic nanostructures. Here we employ self-assembly based on the DNA origami technique for accurate positioning of individual gold nanoparticles. Our innovative design leads to strong plasmonic coupling between two 40 nm gold nanoparticles reproducibly held with gaps of 3.3 ± 1 nm. This is confirmed through far field scattering measurements on individual dimers which reveal a significant red shift in the plasmonic resonance peaks, consistent with the high dielectric environment due to the surrounding DNA. We use surface-enhanced Raman scattering (SERS) to demonstrate local field enhancements of several orders of magnitude through detection of a small number of dye molecules as well as short single-stranded DNA oligonucleotides. This demonstrates that DNA origami is a powerful tool for the high-yield creation of SERS-active nanoparticle assemblies with reliable sub-5 nm gap sizes.

Single Protein Molecule Detection by Glass Nanopores

Nanopores can be used to detect and analyze single molecules in solution. We have used glass nanopores made by laser-assisted capillary-pulling, as a high-throughput and low cost method, to detect a range of label-free proteins: lysozyme, avidin, IgG, β-lactoglobulin, ovalbumin, bovine serum albumin (BSA), and β-galactosidase in solution. Furthermore, we show for the first time solid state nanopore measurements of mammalian prion protein, which in its abnormal form is associated with transmissible spongiform encephalopathies. Our approach provides a basis for protein characterization and the study of protein conformational diseases by nanopore detection.

Lipid-bilayer-spanning DNA nanopores with a bifunctional porphyrin anchor

Holding tight: An artificial membrane nanopore assembled from DNA oligonucleotides carries porphyrin tags (red), which anchor the nanostructure into the lipid bilayer. The porphyrin moieties also act as fluorescent dyes to aid the microscopic visualization of the DNA nanopore.

Multiplexed ionic current sensing with glass nanopores

We report a method for simultaneous ionic current measurements of single molecules across up to 16 solid state nanopore channels. Each device, costing less than

Studying DNA translocation in nanocapillaries using single molecule fluorescence

20, contains 16 glass nanopores made by laser assisted capillary pulling. We demonstrate simultaneous multichannel detection of double stranded DNA and trapping of DNA origami nanostructures to form hybrid nanopores.

Voltage-dependent properties of DNA origami nanopores.

We demonstrate simultaneous measurements of DNA translocation into glass nanopores using ionic current detection and fluorescent imaging. We verify the correspondence between the passage of a single DNA molecule through the nanopore and the accompanying characteristic ionic current blockage. By tracking the motion of individual DNA molecules in the nanocapillary perpendicular to the optical axis and using a model, we can extract an effective mobility constant for DNA in our geometry under high electric fields.

Electroosmotic flow reversal outside glass nanopores.

We show DNA origami nanopores that respond to high voltages by a change in conformation on glass nanocapillaries. Our DNA origami nanopores are voltage sensitive as two distinct states are found as a function of the applied voltage. We suggest that the origin of these states is a mechanical distortion of the DNA origami. A simple model predicts the voltage dependence of the structural change. We show that our responsive DNA origami nanopores can be used to lower the frequency of DNA translocation by 1 order of magnitude.

DNA origami based assembly of gold nanoparticle dimers for SERS detection

We report observations of a striking reversal in the direction of electroosmotic flow (EOF) outside a conical glass nanopore as a function of salt concentration. At high ionic strengths (>100 mM), we observe EOF in the expected direction as predicted by classical electrokinetic theory, while at low salt concentrations (<1 mM) the direction of the flow is reversed. The critical crossover salt concentration depends on the pore diameter. Finite-element simulations indicate a competition between the EOF generated from the inner and outer walls of the pore, which drives flows in opposite directions. We have developed a simple analytical model which reveals that, as the salt concentration is reduced, the flow rates inside the pore are geometrically constrained, whereas there is no such limit for flows outside the pore. This model captures all of the essential physics of the system and explains the observed data, highlighting the key role the external environment plays in determining the overall electroosmotic behavior.

Combining Fluorescence Imaging and Ionic Current Detection in Nanocapillaries

Plasmonic sensors are extremely promising candidates for label-free single molecule analysis but require exquisite control over the physical arrangement of metallic nanostructures. We employ self-assembly based on the DNA origami technique for accurate positioning of individual 40 nm gold nanoparticles with gaps of 3.3±1 nm. This is probed through far field scattering measurements on individual dimers. This plasmonic coupling allows us to use surface enhanced Raman scattering (SERS) to detect a small number of dye molecules as well as short single-stranded DNA oligonucleotides in the vicinity of the dimers. This demonstrates that DNA origami is a powerful tool with great potential for a wide variety of biosensing and single-molecule applications.

DNA origami nanopores for controlling DNA translocation.

We present here recent data showing simultaneous imaging and ionic current detection of DNA fluorescently labeled with SYTOX Orange. Using a fast Electron Multiplying CCD (EMCCD) camera, we have verified the link between ionic current translocation events and the movement of the DNA into the nanocapillary. Further work will focus on elucidating the nature of so-called ‘folding’ DNA events as well as studies on the balance of electrophoresis and electroosmosis in nanocapillaries.