Despite demonstrating exciting potential for applications such as drug delivery and biosensing, the development of nanodevices for practical applications and broader use in research and education are still hindered by the time, effort, and cost associated with DNA origami fabrication. Simple and robust methods to perform and scale the DNA origami self-assembly process are critical to facilitate broader use and translation to industrial or clinical applications. We report a simple approach to fold DNA origami nanostructures that is fast, robust, and scalable. We demonstrate fabrication at scales approximately 100–1,500-fold higher than typical scales. We further demonstrate an approach we termed low-cost efficient annealing (LEAN) self-assembly involving initial heating at 65 °C for 10 min, then annealing at 51 °C for 2 h, followed by brief quenching at 4 °C that leads to effective assembly of a range of DNA origami structures tested. In contrast to other methods for scaling DNA origami assembly, this approach can be carried out using cheap and widely available equipment (e.g., hot plates, water baths, and laboratory burners) and uses standard recipes and materials so is readily applied to any existing or new DNA origami designs. We envision these methods can facilitate device development for commercial applications and facilitate broader use of DNA origami in research and education.

The ability to design and control DNA nanodevices with programmed conformational changes has established a foundation for molecular-scale robotics with applications in nanomanufacturing, drug delivery, and controlling enzymatic reactions. The most commonly used approach for actuating these devices, DNA binding and strand displacement, allows devices to respond to molecules in solution, but this approach is limited to response times of minutes or greater. Recent advances have enabled electrical and magnetic control of DNA structures with sub-second response times, but these methods utilize external components with additional fabrication requirements. Here, we present a simple and broadly applicable actuation method based on the avidity of many weak base-pairing interactions that respond to changes in local ionic conditions to drive large-scale conformational transitions in devices on sub-second time scales. To demonstrate such ion-mediated actuation, we modified a DNA origami hinge with short, weakly complementary single-stranded DNA overhangs, whose hybridization is sensitive to cation concentrations in solution. We triggered conformational changes with several different types of ions including mono-, di-, and trivalent ions and also illustrated the ability to engineer the actuation response with design parameters such as number and length of DNA overhangs and hinge torsional stiffness. We developed a statistical mechanical model that agrees with experimental data, enabling effective interpretation and future design of ion-induced actuation. Single-molecule Förster resonance energy-transfer measurements revealed that closing and opening transitions occur on the millisecond time scale, and these transitions can be repeated with time resolution on the scale of one second. Our results advance capabilities for rapid control of DNA nanodevices, expand the range of triggering mechanisms, and demonstrate DNA nanomachines with tunable analog responses to the local environment.

Biological movement and processes are ultimately driven by the interactions of singular biomolecules. However, only in the past few decades have techniques and technologies (e.g. optical trapping, atomic force microscopy, and magnetic tweezers) been developed capable of probing these interactions. Experiments utilizing these methodologies are often encumbered by high costs, low-throughput, and reliant upon user skill or expertise. The focus of this work is the development, characterization, validation, and application of a nanoscale device designed explicitly for fluorescence-based single molecule force spectroscopy. This device allows for force dependent measurements of single molecular interaction kinetics to be performed on 100s of interactions simultaneously using a basic laboratory fluorescence microscope. The nanostructure comprises a stiff platform that contains functionalized attachment points for two biomolecules (a receptor and a ligand), a flexible single stranded DNA (ssDNA) linker that as an entropic spring, and fluorescent molecules to facilitate readout of the binding interaction between the biomolecules under study via a FRET interaction. This device will enable the investigation of the kinetics and mechanical stability of biomolecular interactions and singular biomolecules in a highly parallel fashion. We have constructed the device and performed proof of principle experiments probing DNA base-pairing interactions. We have characterized the device and have shown it is able to facilitate and rupture these interactions with estimated forces ranging from ∼1-20pN. A single molecule FRET assay has been developed and is initially being used to measure the kinetics of previously characterized DNA interactions to verify the behavior of the device. After verifying the functionality, this device will be used to probe the receptor-ligand kinetics of DNA-protein interactions.

DNA-origami nanodevices are embedded on the membrane of two cells by Jonathan W. Song, Carlos E. Castro, and co-workers in article number 1703632. The devices are anchored to the cell via a DNA base-pairing attachment to a cholesterol-labeled DNA strand embedded in the membrane. The two pairs of nanostructures (shown here on the right) mediate adhesion between the cells via a sequence-specific DNA binding interaction.

A specific and reversible method is reported to engineer cell‐membrane function by embedding DNA‐origami nanodevices onto the cell surface. Robust membrane functionalization across epithelial, mesenchymal, and nonadherent immune cells is achieved with DNA nanoplatforms that enable functions including the construction of higher‐order DNA assemblies at the cell surface and programed cell–cell adhesion between homotypic and heterotypic cells via sequence‐specific DNA hybridization. It is anticipated that integration of DNA‐origami nanodevices can transform the cell membrane into an engineered material that can mimic, manipulate, and measure biophysical and biochemical function within the plasma membrane of living cells.

SmallVolume 12, Issue 3 p. 307-307 FrontispieceFree Access DNA Origami: Daunorubicin-Loaded DNA Origami Nanostructures Circumvent Drug-Resistance Mechanisms in a Leukemia Model (Small 3/2016) Patrick D. Halley, Patrick D. Halley Department of Mechanical and Aerospace Engineering, College of Engineering, The Ohio State University, Columbus, OH, 43210 USA Department of Chemical and Biomolecular Engineering, College of Engineering, The Ohio State University, Columbus, OH, 43210 USASearch for more papers by this authorChristopher R. Lucas, Christopher R. Lucas Department of Mechanical and Aerospace Engineering, College of Engineering, The Ohio State University, Columbus, OH, 43210 USASearch for more papers by this authorEmily M. McWilliams, Emily M. McWilliams Biomedical Sciences Graduate Program, Department of Internal Medicine, College of Medicine, The Ohio State University, Columbus, OH, 43210 USASearch for more papers by this authorMatthew J. Webber, Matthew J. Webber Biophysics Graduate Program, The Ohio State University, Columbus, OH, 43210 USASearch for more papers by this authorRandy A. Patton, Randy A. Patton Department of Mechanical and Aerospace Engineering, College of Engineering, The Ohio State University, Columbus, OH, 43210 USASearch for more papers by this authorComert. Kural, Comert. Kural Biophysics Graduate Program, Department of Physics, The Ohio State University, Columbus, OH, 43210 USASearch for more papers by this authorDavid M. Lucas, David M. Lucas Department of Internal Medicine, College of Medicine, College of Pharmacy, The Ohio State University, Columbus, OH, 43210 USASearch for more papers by this authorJohn C. Byrd, John C. Byrd Department of Internal Medicine, College of Medicine, College of Pharmacy, The Ohio State University, Columbus, OH, 43210 USASearch for more papers by this authorCarlos E. Castro, Corresponding Author Carlos E. Castro Department of Mechanical and Aerospace Engineering, College of Engineering, Biophysics Graduate Program, The Ohio State University, Columbus, OH, 43210 USASearch for more papers by this author Patrick D. Halley, Patrick D. Halley Department of Mechanical and Aerospace Engineering, College of Engineering, The Ohio State University, Columbus, OH, 43210 USA Department of Chemical and Biomolecular Engineering, College of Engineering, The Ohio State University, Columbus, OH, 43210 USASearch for more papers by this authorChristopher R. Lucas, Christopher R. Lucas Department of Mechanical and Aerospace Engineering, College of Engineering, The Ohio State University, Columbus, OH, 43210 USASearch for more papers by this authorEmily M. McWilliams, Emily M. McWilliams Biomedical Sciences Graduate Program, Department of Internal Medicine, College of Medicine, The Ohio State University, Columbus, OH, 43210 USASearch for more papers by this authorMatthew J. Webber, Matthew J. Webber Biophysics Graduate Program, The Ohio State University, Columbus, OH, 43210 USASearch for more papers by this authorRandy A. Patton, Randy A. Patton Department of Mechanical and Aerospace Engineering, College of Engineering, The Ohio State University, Columbus, OH, 43210 USASearch for more papers by this authorComert. Kural, Comert. Kural Biophysics Graduate Program, Department of Physics, The Ohio State University, Columbus, OH, 43210 USASearch for more papers by this authorDavid M. Lucas, David M. Lucas Department of Internal Medicine, College of Medicine, College of Pharmacy, The Ohio State University, Columbus, OH, 43210 USASearch for more papers by this authorJohn C. Byrd, John C. Byrd Department of Internal Medicine, College of Medicine, College of Pharmacy, The Ohio State University, Columbus, OH, 43210 USASearch for more papers by this authorCarlos E. Castro, Corresponding Author Carlos E. Castro Department of Mechanical and Aerospace Engineering, College of Engineering, Biophysics Graduate Program, The Ohio State University, Columbus, OH, 43210 USASearch for more papers by this author First published: 14 January 2016 https://doi.org/10.1002/smll.201670014Citations: 5AboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Share a linkShare onFacebookTwitterLinkedInRedditWechat Graphical Abstract DNA origami "Trojan horse" nanostructures provide an effective delivery approach to circumvent drug resistance in leukemia cells. These 100 nanometer delivery vehicles, described on page 308 by C. E. Castro and co-workers, are easily fabricated in 10 min and can be precisely loaded with commonly used anthracycline chemotherapeutic drugs. Drug-loaded DNA origami nanostructures enter cells via endocytosis, allowing the drug to bypass defenses in the cell membrane that are effective against free drug. This approach leads to larger amounts of drug in cells, which is later released due to decreasing pH and enzymatic degradation, ultimately improving drug efficacy. Citing Literature Volume12, Issue3January 20, 2016Pages 307-307 RelatedInformation

Many cancers show primary or acquired drug resistance due to the overexpression of efflux pumps. A novel mechanism to circumvent this is to integrate drugs, such as anthracycline antibiotics, with nanoparticle delivery vehicles that can bypass intrinsic tumor drug-resistance mechanisms. DNA nanoparticles serve as an efficient binding platform for intercalating drugs (e.g., anthracyclines doxorubicin and daunorubicin, which are widely used to treat acute leukemias) and enable precise structure design and chemical modifications, for example, for incorporating targeting capabilities. Here, DNA nanostructures are utilized to circumvent daunorubicin drug resistance at clinically relevant doses in a leukemia cell line model. The fabrication of a rod-like DNA origami drug carrier is reported that can be controllably loaded with daunorubicin. It is further directly verified that nanostructure-mediated daunorubicin delivery leads to increased drug entry and retention in cells relative to free daunorubicin at equal concentrations, which yields significantly enhanced drug efficacy. Our results indicate that DNA origami nanostructures can circumvent efflux-pump-mediated drug resistance in leukemia cells at clinically relevant drug concentrations and provide a robust DNA nanostructure design that could be implemented in a wide range of cellular applications due to its remarkably fast self-assembly (≈5 min) and excellent stability in cell culture conditions.

Structural DNA nanotechnology is a rapidly emerging field with immense potential for applications such as single molecule sensing, drug delivery, and manipulating molecular components. Major advances in the last decade have enabled the precise design and fabrication of DNA nanostructures with unprecedented geometric complexity; however, relative to natural biomolecular machines, the functional scope of DNA nanotechnology is limited by an inability to design dynamic mechanical behavior such as complex motion, conformational dynamics, or force generation. Inspired by approaches used in macroscopic machine design, we have recently developed methods to design structures with controllable 1D, 2D, or 3D motion. Moving beyond design of geometry or motion paths, we have recently developed structures with multiple thermally accessible states separated by well-defined energy barriers with tunable height. In particular, we have design a two-state device that transitions between compact and open states driven by thermal energy. We directly measured the dynamic behavior via single molecule fluorescence experiments, which show both the equilibrium distribution of states and the kinetics of switching between states can be tuned via design parameters that regulate the relative configurational space, or equivalently the entropy change, between states. Finally, we show that these dynamic structures can be used to probe local molecular scale forces, specifically depletion forces that result from entropic interactions with a crowding reagent. This work is the first demonstration of an ability to control the kinetics of DNA origami nanostructures down to the second scale and establishes a foundation for exploiting dynamic behavior of these devices to study physical interactions at the molecular scale.

In this manuscript, we investigate a new methodology for increasing the spectral purity of the second-harmonic output of an injection-seeded, frequency-doubled, Q-switched Nd:YAG laser operating near 532 nm. Specifically, tunable Fabry–Perot etalons (FPEs) are used as ultra-narrowband spectral filters, transmitting the desired single-mode output, while filtering out a significant portion of the broadband pedestal characteristic of injection-seeded lasers. A specific emphasis is placed on the design and optimization of the FPEs in the context of filtered Rayleigh scattering (FRS) measurements and how their utilization results in substantial increases in spectral purity, realizable attenuation of unwanted scattering, and applications in environments with high particulate levels. Experimental results show an increase in laser spectral purity of more than one order-of-magnitude (from 0.99997 to 0.999998) when using FPE filters, which led to a two-order-of-magnitude increase in achievable attenuation of laser light passing through a molecular iodine filter. The utility of the FPE-based spectral filtering of the pulsed Nd:YAG output for 2D FRS imaging was demonstrated in turbulent, isothermal gas-phase jets, seeded with varying levels of non-evaporating droplets with particle volume fractions (F Vp) ranging from ~5 to >60 parts-per-million (ppm). After implementation of an optimized air-spaced FPE in the 532-nm output, no particle scattering was observed (based on visual and statistical analysis), even for the highest seed case (F Vp ~ 60 ppm), and the gas-phase Rayleigh–Brillouin signals were collected without interference from the flowfield particulate. The current results suggest that the implementation of properly specified FPEs allows FRS to be applied in environments with high flowfield particulate levels; levels are well beyond what have been suitable for previous FRS measurements.

In this manuscript, we investigate the role of seed laser power on the spectral purity of an injection-seeded, frequency-doubled, Q-switched Nd:YAG laser operating near 532 nm. A specific emphasis is placed on examining how the seed laser power and the corresponding spectral purity affects molecular iodine-filtered Rayleigh scattering (FRS) measurements in terms of realizable attenuation of unwanted scattering. Results from a set of detailed calculations demonstrate a logarithmic relationship between spectral purity and the effective optical density of the I (2) lines, that is, for every order-of-magnitude decrease in spectral purity, there is a corresponding order-of-magnitude decrease in achievable optical density. As one example, a decrease in spectral purity from 1 to 0.999999 results in an increase in the minimum transmission of > 10(3). We directly measured the optical density of strong I (2) absorption lines for a series of injection seed powers ranging from 1 to 15 mW. For this range of circulating seed power, measured optical densities increased from 4 to 5.2 and the corresponding spectral purities were estimated as increasing from 0.9999 to 0.999992. The relationship between measured spectral purity and seed power agreed well with previous theoretical scaling. The current results imply that one potential methodology of achieving greater suppression of unwanted scattering when using molecular iodine-filtered Rayleigh scattering (FRS) near 532 nm is to increase the power of the seed laser.

We report new results using laser-based imaging techniques (CH2O PLIF and Mie scattering) to understand the structure of turbulent dimethyl ether-based flames in comparison to well-studied methane-based flames. Specifically, the paper introduces a new set of potential target flames using DME fuel. The new DME-based flames are formulated with the specific intention of directly comparing non-intrusive laser-based measurements with the DLR A and B flames, which serve as target cases within the TNF workshop. The new DME-based flames are operated at equivalent Reynolds numbers and have an identical stoichiometric mixture fraction as the DLR flames, thus attempting to isolate kinetic effects and turbulence-chemistry interaction from flowfield and mixing effects. In this paper, we examine the CH2O field using planar laser-induced fluorescence (PLIF) and laser Mie scattering of fuel–seeded oil droplets which yield an approximate marker of the 700–800K isotherms. Results from laminar flame calculations indicate that large amounts of CH2O are formed in DME-based flames as compared to CH4-based flames due to very rapid DME pyrolysis. Additionally, the calculations suggest significant differences in the topology of the CH2O distribution. Consistent with the laminar flame calculations, the measured CH2O PLIF signals from the turbulent DME-based flames are approximately two orders of magnitude higher than the DLR flames. Furthermore, instantaneous, average, and RMS images from the turbulent flames are used to highlight qualitative differences in the CH2O field for the DME-based flames in comparison with the methane-based DLR flames.

In this paper we will describe recent advances made in our laboratory towards the development, validation, and application of a filtered Rayleigh Scattering technique for measurements of gas-phase concentration fields in the presence of liquid-phase droplets. Preliminary studies of the maximum level of droplet scattering attenuation attainable utilizing an injection-seeded, Q-Switched Nd:YAG laser in conjunction with a molecular iodine filter will be presented. These results demonstrate the correlation of the spectral purity of the laser with the effectiveness of the molecular filter in suppressing particle (Mie) scattering. Improvements in spectral purity and the maximum Mie scattering rejection as a result of implementing an external etalon into the optical path of the laser output will be demonstrated. Initial validation studies of the FRS technique also are presented in which gas-phase information is extracted from a droplet-laden turbulent jet. Comparisons of suppression capabilities with and without the external etalon are presented, allowing for an assessment gains offered in implementing the etalon. Finally, we present initial results from an evaporating spray jet of acetone into air.

In this study, we describe the development of two-dimensional, high repetition-rate (10-kHz) Rayleigh scattering imaging as applied to turbulent flows. In particular, we report what we believe to be the first sets of high-speed 2D Rayleigh scattering images in turbulent non-reacting jets, yielding temporally correlated image sequences of the instantaneous mixture fraction field. Results are presented for turbulent jets of propane issuing into a low-speed co-flow of air at jet-exit Reynolds numbers of 10,000, 15,000, and 30,000 at various axial positions downstream of the jet exit. The quantitative high-speed mixture fraction measurements are facilitated by the use of a calibrated, un-intensified, high-resolution CMOS camera in conjunction with a unique high-energy, high-repetition rate pulse-burst laser system (PBLS) at Ohio State, which yields output energies of ∼200 mJ/pulse at 532 nm with 100-μs laser pulse spacing. The quality, accuracy, and resolution of the imaging system and the resulting image sets are assessed by (1) comparing the mean mixture fraction results to known scaling laws for turbulent jets, (2) comparing instantaneous images/mixture fraction profiles acquired simultaneously with the high-speed CMOS camera and a well-characterized, high-quantum efficiency CCD camera, and (3) comparing statistical quantities such as the probability density function of the mixture fraction results using the high-speed CMOS camera and the CCD camera. Results indicate accurate mixture fraction measurements and a high potential for accurately measuring mixture fraction gradients in both time and space.

Nitric oxide planar laser-induced fluorescence (NO PLIF) imaging is demonstrated at a 10-kHz repetition rate in the Calspan University at Buffalo Research Center’s (CUBRC) 48-inch Mach 9 hypervelocity shock tunnel using a pulse burst laser–based high frame rate imaging system. Sequences of up to ten images are obtained internal to a supersonic combustor model, located within the shock tunnel, during a single ~10-millisecond duration run of the ground test facility. Comparison with a CFD simulation shows good overall qualitative agreement in the jet penetration and spreading observed with an average of forty individual PLIF images obtained during several facility runs.

In this manuscript, we demonstrate high-speed (10-kHz-acquisition rate) planar laser-induced fluorescence (PLIF) imaging of formaldehyde (CH2O) in turbulent non-premixed flames. Using the unique pulse-burst laser system (PBLS) at Ohio State University, high-energy laser pulses (∼100 mJ/pulse) at 355 nm with 100 μs pulse separation are generated and used to measure the time-varying CH2O distributions in attached and lifted methane-based turbulent flames. By taking advantage of the tunable, narrow spectral linewidth of the PBLS at 355 nm, the laser output can be frequency-tuned and adjusted to overlap with absorption “peaks” within the tail of the A–X transition of CH2O near 355 nm, thus increasing the acquired signal by as much as a factor of three. The reported signal-to-noise ratio (SNR) exceeds 55, which represents one of the highest SNR reported to date for kilohertz-rate imaging of scalars for comparable spatial resolution. Potential applications and pairings with other diagnostic approaches for high-speed reaction rate and multi-scalar imaging also are discussed.

In this manuscript, we describe the development of two-dimensional, high-repetition-rate (10-kHz) Rayleigh scattering imaging as applied to turbulent combustion environments. In particular, we report what we believe to be the first sets of high-speed planar Rayleigh scattering images in turbulent non-premixed flames, yielding temporally correlated image sequences of the instantaneous temperature field. Sample results are presented for the well-characterized DLR flames A and B (CH4/H2/N2) at Reynolds numbers of 15,200 and 22,800 at various axial positions downstream of the jet exit. The measurements are facilitated by the use of a user-calibrated, intensified, high-resolution CMOS camera in conjunction with a unique high-energy, high-repetition-rate pulse-burst laser system (PBLS) at Ohio State University, which yields output energies up to 200 mJ/pulse at 532 nm with 100-μs laser pulse spacing. The spatial and temporal resolution of the imaging system and acquired images are compared to the finest spatial and temporal scales expected within the turbulent flames. One of the most important features of the PBLS is the ability to readily change the pulse-to-pulse spacing as the required temporal resolution necessitates it. The quality and accuracy of the high-speed temperature imaging results are assessed by comparing derived statistics (mean and standard deviation) to that of previously reported point-based reference data acquired at Sandia National Laboratories and available within the TNF workshop. Good agreement between the two data sets is obtained providing an initial indication of quantitative nature of the planar, kHz-rate temperature imaging results.

In this paper we will describe recent advances made in our laboratory in the development of both high-speed mixture fraction ([) imaging and simultaneous mixture fraction and velocity imaging in turbulent nonreacting jets. High-repetition rate mixture fraction imaging has been developed in turbulent jets using planar Rayleigh scattering imaging, with the goal of extending this measurement capability to include simultaneous high-speed planar velocity measurements using particle imaging velocimetry (PIV). In this respect, conventional Rayleigh scattering imaging will be replaced with Filtered Rayleigh scattering (FRS), so that [ [ [ [ can be measured in the presence of the PIV seed particles without interference. In this paper, we will report a successful demonstration of high-speed (10 kHz) mixture fraction imaging using Rayleigh scattering and progress towards simultaneous mixture fraction and velocity imaging using FRS/Mie scattering at low (10 Hz) repetition rates. Future work entails merging the two independent work paths to enable a simultaneous high-speed mixture fraction/velocity measurement capability.

In this paper we will describe recent advances made in our laboratory in the development of high-repetition-rate Rayleigh and Raman scattering imaging capabilities. High-repetition-rate 1D Raman and 2D Rayleigh scattering imaging capabilities are being developed to image the time-varying mixture fraction and temperature fields in turbulent non-reacting and reacting flows. Initial results using a custom pulse-burst laser system at Ohio State University have demonstrated the ability to capture ten sequential 2D Rayleigh scattering images at a repetition rate of 10 kHz in both turbulent non-reacting jets and non-premixed jet-flames with pulse energies approaching 200 mJ at 532 nm. This paper will also describe the development and pending application of a new higher-energy, long-duration, next-generation burst-mode laser system and the use of higher resolution cameras for high-speed Raman/Rayleigh imaging.