Hyunmin Yi

Detecting hybridization of DNA by highly sensitive evanescent field etched core fiber Bragg grating sensors

Highly sensitive fiber Bragg grating sensors were developed by etching away the cladding and part of the core of the fiber and detecting the change of Bragg wavelength due to the change of index of the surrounding medium. A sensitivity of 1394 nm/riu was achieved when the diameter of the grating core was 3.4 /spl mu/m and the index of the surrounding medium was close to the index of the core of the fiber. Assuming a detectable spectral resolution of 0.01 nm realized in the experiment, the sensor achieves a minimum detectable index resolution of 7.2/spl times/10/sup -6/. Higher sensitivity at lower surrounding index was achieved by using higher order modes excited in the Bragg grating region. The use of the fiber Bragg grating sensor was further investigated to detect hybridization of DNA. Single stranded DNA oligonucleotide probes of 20 bases were immobilized on the surface of the fiber grating using relatively common glutarahyldehyde chemistry. Hybridization of complimentary target single strand DNA oligonucleotide was monitored in situ and successfully detected. The demonstrated fiber Bragg grating sensors provide an elegant method to monitor biological changes in an in situ manner, and provide temporal information in a single experiment.

Simple, Readily Controllable Palladium Nanoparticle Formation on Surface-Assembled Viral Nanotemplates

Transition-metal nanoparticles possess unique size-dependent optical, electronic, and catalytic properties on the nanoscale, which differ significantly from their bulk properties. In particular, palladium (Pd) nanoparticles have properties applicable to a wide range of applications in catalysis and electronics. However, predictable and controllable nanoparticle synthesis remains challenging because of harsh reaction conditions, artifacts from capping agents, and unpredictable growth. Biological supramolecules offer attractive templates for nanoparticle synthesis because of their precise structure and size. In this article, we demonstrate simple, controllable Pd nanoparticle synthesis on surface-assembled viral nanotemplates. Specifically, we exploit precisely spaced thiol functionalities of genetically modified tobacco mosaic virus (TMV1cys) for facile surface assembly and readily controllable Pd nanoparticle synthesis via simple electroless deposition under mild aqueous conditions. Atomic force microscopy (AFM) studies clearly show tunable surface assembly and Pd nanoparticle formation preferentially on the TMV1cys templates. Grazing incidence small-angle X-ray scattering (GISAXS) further provided an accurate and statistically meaningful route by which to investigate the broad size ranges and uniformity of the Pd nanoparticles formed on TMV templates by simply tuning the reducer concentration. We believe that our viral-templated bottom-up approach to tunable Pd nanoparticle formation combined with the first in-depth characterization via GISAXS represents a major advancement toward exploiting viral templates for facile nanomaterials/device fabrication. We envision that our strategy can be extended to a wide range of applications, including uniform nanostructure and nanocatalyst synthesis.

Viral-templated palladium nanocatalysts for Suzuki coupling reaction

We demonstrate and thoroughly examine tobacco mosaic virus (TMV)-templated palladium (Pd) nanocatalysts for the ligand-free Suzuki coupling reaction under mild conditions. The surface-assembled TMV templates allow for facile catalyst synthesis under mild aqueous conditions that leads to high Pd surface loading and stability. Further, the chip-based format enables simple catalyst separation and reuse as well as facile product recovery. Reaction condition studies demonstrated that the solvent ratio played an important role in the selectivity of the Suzuki reaction, and that a higher water/acetonitrile ratio significantly facilitated the cross-coupling pathway. We envision that our viral template-based bottom-up assembly approach can be readily extended to other biotemplates, metal catalysts and organic reaction systems.

Fabrication of Uniform DNA-Conjugated Hydrogel Microparticles via Replica Molding for Facile Nucleic Acid Hybridization Assays

We identify and investigate several critical parameters in the fabrication of single-stranded DNA conjugated poly(ethylene glycol) (PEG) microparticles based on replica molding (RM) for highly uniform and robust nucleic acid hybridization assays. The effects of PEG-diacrylate, probe DNA, and photoinitiator concentrations on the overall fluorescence and target DNA penetration depth upon hybridization are examined. Fluorescence and confocal microscopy results illustrate high conjugation capacity of the probe and target DNA, femtomole sensitivity, and sequence specificity. Combined, these findings demonstrate a significant step toward simple, robust, and scalable procedures to manufacture highly uniform and high-capacity hybridization assay particles in a well-controlled manner by exploiting many advantages that the batch processing-based RM technique offers. We envision that the results presented here may be readily applied to rapid and high-throughput hybridization assays for a wide variety of applications in bioprocess monitoring, food safety, and biological threat detection.

Mechano-transduction of DNA hybridization and dopamine oxidation through electrodeposited chitosan network.

While microcantilevers offer exciting opportunities for mechano-detection, they often suffer from limitations in either sensitivity or selectivity. To address these limitations, we electrodeposited a chitosan film onto a cantilever surface and mechano-transduced detection events through the chitosan network. Our first demonstration was the detection of nucleic acid hybridization. In this instance, we electrodeposited the chitosan film onto the cantilever, biofunctionalized the film with oligonucleotide probe, and detected target DNA hybridization by cantilever bending in solution (static mode) or resonant frequency shifts in air (dynamic mode). In both detection modes, we observed a two-order of magnitude increase in sensitivity compared to values reported in literature for DNA immobilized on self-assembled monolayers. In our second demonstration, we coupled electrochemical and mechanical modes to selectively detect the neurotransmitter dopamine. A chitosan-coated cantilever was biased to electrochemically oxidize dopamine solution. Dopamines oxidation products react with the chitosan film and create a tensile stress of approximately 1.7 MPa, causing substantial cantilever bending. A control experiment was performed with ascorbic acid solution. It was shown that the electrochemical oxidation of ascorbic acid does not lead to reactions with chitosan and does not change cantilever bending. These results suggest that chitosan can confer increased sensitivity and selectivity to microcantilever sensors.

A Facile Synthesis–Fabrication Strategy for Integration of Catalytically Active Viral-Palladium Nanostructures into Polymeric Hydrogel Microparticles via Replica Molding

The synthesis of small, uniform, well-dispersed and active Pd nanocatalysts under mild conditions in a predictable and controlled manner is an unmet challenge. Viral nanomaterials are attractive biotemplates for the controlled synthesis of nanoparticles due to their well-defined and monodisperse structure along with abundant surface functionalities. Here, we demonstrate spontaneous formation of small (1-2 nm), uniform and highly crystalline palladium (Pd) nanoparticles along genetically modified tobacco mosaic virus (TMV1cys) biotemplates without external reducing agents. The ratio between TMV and Pd precursor plays an important role in the exclusive formation of well-dispersed Pd nanoparticles along TMV biotemplates. The as-prepared Pd-TMV complexes are then integrated into the poly(ethylene glycol) (PEG)-based microparticles via replica molding (RM) technique in a simple, robust and highly reproducible manner. High catalytic activity, recyclability and stability of the hybrid Pd-TMV-PEG microparticles are further demonstrated through dichromate reduction as a model reaction. Taken together, these findings demonstrate a significant step toward simple, robust, and scalable synthesis and fabrication of efficient biotemplate-supported Pd nanocatalysts in readily deployable polymeric scaffolds with high capacity in a controlled manner.

Fabrication of chitosan-poly(ethylene glycol) hybrid hydrogel microparticles via replica molding and its application toward facile conjugation of biomolecules.

We demonstrate a facile scheme to fabricate nonspherical chitosan-poly(ethylene glycol) (PEG) microparticle platforms for conjugation of biomolecules with high surface density. Specifically, we show that PEG microparticles containing short chitosan oligomers are readily fabricated via replica molding (RM). Fluorescence and FTIR microscopy results illustrate that these chitosan moieties are incorporated with PEG networks in a stable manner while retaining chemical reactivity toward amine-reactive chemistries. The chitosan-PEG particles are then conjugated with single-stranded (ss) DNAs via Cu-free click chemistry. Fluorescence and confocal microscopy results show facile conjugation of biomolecules with the chitosan-PEG particles under mild conditions with high selectivity. These ssDNA-conjugated chitosan-PEG particles are then enlisted to assemble tobacco mosaic virus (TMV) via nucleic acid hybridization as an example of orientationally controlled conjugation of supramolecular targets. Results clearly show controllable TMV assembly with high surface density, indicating high surface DNA density on the particles. Combined, these results demonstrate a facile fabrication-conjugation scheme for robust biomolecular conjugation or assembly platforms. We expect that our approach can be enlisted in a wide array of biomolecular targets and applications.

Facile Strategy for Protein Conjugation with Chitosan-Poly(ethylene glycol) Hybrid Microparticle Platforms via Strain-Promoted Alkyne–Azide Cycloaddition (SPAAC) Reaction

We demonstrate a facile fabrication-conjugation scheme for protein-conjugated biosensing platforms. Specifically, we utilize a chitosan-poly(ethylene glycol) hybrid system to fabricate highly uniform and chemically reactive microparticle platforms via simple replica molding. Strain-promoted alkyne-azide cycloaddition (SPAAC) reaction between azide-modified proteins and microparticles activated with strain-promoted cyclooctynes allows tunable protein conjugation under mild reaction conditions. Upon conjugation of a model red fluorescent protein, fluorescence and confocal micrographs show selective protein conjugation near the particle surfaces as well as long-term stability of the conjugation scheme. Fluorescence and AFM results upon conjugation with varying protein concentrations indicate controllable protein conjugation. Examination of protein-particle conjugation kinetics shows multiple reaction regimes; rapid initial, intermediate, and steady final stage. Lastly, we demonstrate antibody conjugation with the particles and selective and rapid target protein capture with antibody-conjugated particles. Combined, these results illustrate a facile fabrication-conjugation scheme for robust protein-conjugated platforms that can be readily enlisted in various protein sensing applications.

In Situ Small-Angle X-ray Scattering Analysis of Palladium Nanoparticle Growth on Tobacco Mosaic Virus Nanotemplates

We present an examination of palladium (Pd) nanoparticle growth on genetically modified tobacco mosaic virus (TMV1cys) nanotemplates via in situ small-angle X-ray scattering (SAXS). Specifically, we examine the role of the TMV1cys templates in Pd nanoparticle formation through the electroless reduction of Pd precursor by a chemical reducing agent as compared to identical conditions in the absence of the TMV1cys templates. We show that in the presence of TMV1cys, the viral nanotemplates provide preferential growth sites for Pd nanoparticle formation, as no measurable Pd particle growth was observed in the bulk solution. In situ SAXS confirmed that particle formation was due to the rapid adsorption of Pd atoms onto the TMV1cys templates at the very early stage of mixing, rather than adsorption of particles formed in the bulk solution. Importantly, Pd nanoparticles were significantly smaller and more uniform as compared to particle formation in the absence of TMV1cys. The Pd nanoparticle coating density was tunable based on Pd precursor concentration. Finally, we show that Pd particle growth on the TMV1cys templates was highly rapid, and complete within 33 s for most samples, in contrast to slower Pd particle growth in the absence of TMV templates. We envision that the results presented here will be valuable in furthering the fundamental understanding of the role of viral nanotemplates in particle formation, as well as of their utility in a wide range of applications.

An integrated approach for enhanced protein conjugation and capture with viral nanotemplates and hydrogel microparticle platforms via rapid bioorthogonal reactions.

We demonstrate significantly enhanced protein conjugation and target protein capture capacity by exploiting tobacco mosaic virus (TMV) templates assembled with hydrogel microparticles. Protein conjugation results with a red fluorescent protein R-Phycoerythrin (R-PE) show significantly enhanced protein conjugation capacity of TMV-assembled particles (TMV-particles) compared to planar substrates or hydrogel microparticles. In-depth examination of protein conjugation kinetics via tetrazine (Tz)-trans-cyclooctene (TCO) cycloaddition and strain-promoted alkyne-azide cycloaddition (SPAAC) reaction demonstrates that TMV-particles provide a less hindered environment for protein conjugation. Target protein capture results using an anti-R-PE antibody (R-Ab)-R-PE pair also show substantially improved capture capacity of R-Ab conjugated TMV-particles over R-Ab conjugated hydrogel microparticles. We further demonstrate readily controlled protein and antibody conjugation capacity by simply varying TMV concentrations, which show negligible negative impact of densely assembled TMVs on protein conjugation and capture capacity. Combined, these results illustrate a facile postfabrication protein conjugation approach with TMV templates assembled onto hydrogel microparticles for improved and controlled protein conjugation and sensing platforms. We anticipate that our approach can be readily applied to various protein sensing applications.