Biomedical subjects

Yong-Kweon Kim

Publications and source records attributed to Yong-Kweon Kim.

12 recordsLinked to original sources

PubMed2006

Nanoparticle probes with surface enhanced Raman spectroscopic tags for cellular cancer targeting.

We have developed biocompatible, photostable, and multiplexing-compatible surface-enhanced Raman spectroscopic tagging material (SERS dots) composed of silver nanoparticle-embedded silica spheres and organic Raman labels for cellular cancer targeting in living cells. SERS dots showed linear dependency of Raman signatures on their different amounts, allowing their possibility for the quantification of targets. In addition, the antibody-conjugated SERS dots were successfully applied to the targeting of HER2 and CD10 on cellular membranes and exhibited good specificity. SERS dots demonstrate the potential for high-throughput screening of biomolecules using vibrational information.

Humans↗

PubMed2006

Application of a temperature-controllable microreactor to simple and rapid protein identification using MALDI-TOF MS.

We have carried out a simultaneous thermal denaturation and trypsin digestion of proteins using a temperature-controllable microreactor. This is a simple and rapid sample preparation technique for use before matrix-assisted laser desorption ionization time-of-flight mass spectrometry. In contrast to a conventional sample preparation method, which involves several chemical treatments, our sample preparation was performed using only trypsin digestion with the thermal denaturation of the target protein. Optimization of the reactor operational parameters for trypsin digestion using a temperature-controllable microreactor was carried out. The entire trypsin digestion procedure took about 11 min, and consisted of 1 min for the thermal denaturation of the sample protein (3 microl, 0.2 microM) at 85 degrees C, and 10 min for digestion of the protein at 37 degrees C. The resulting sequence coverage ranged from 24% to 57%, which was sufficient for practical protein identification.

Microchemistry↗

PubMed2006

A method of binding kinetics of a ligand to micropatterned proteins on a microfluidic chip.

A combination of microfluidic protein patterning and quantitative microfluidic handling has been used to analyze the binding kinetics of protein-ligand interactions on the nanoliter scale. The microfluidic handling method employing hydrophobic valving and pneumatic control allowed us to control nanoliter volumes of ligand or protein on a microfluidic chip. A hydrophobic and inert fluorocarbon thin film was patterned on a silicon nitride substrate to prevent non-specific binding on the background. Selectively patterned protein patterns of various sizes were used for quantitative analysis of the kinetic parameters of immobilized proteins on the circular patterns. As a model system, a streptavidin-patterned array of the same-sized pattern, i.e. 150 microm diameter, was used to capture FITC-BSA-biotin present in solution. The fluorescence intensity was well matched with the Langmuir isotherm model results, showing a dissociation constant of 2.43x10(-8)M. Similar streptavidin arrays with different-sized spots, ranging from 50 to 200 microm, showed a consistent dissociation constant of FITC-BSA-biotin with streptavidin pattern. Therefore, the reduction of pattern size of an immobilized protein did not change the dissociation rate of the ligand.

Biosensing Techniques↗

PubMed2005

Bias-free pneumatic sample injection in microchip electrophoresis.

We have developed a new microfluidic chip capable of accurate metering, pneumatic sample injection, and subsequent electrophoretic separation. The pneumatic injection scheme, enabling us to introduce a solution without sampling bias unlike electrokinetic injection, is based upon the hydrophobicity and wettability of channel surfaces. An accurately metered solution of 10 nL could be injected by pneumatic pressure into a hydrophilic separation channel through Y-shaped hydrophobic valves, which consist of polydimethylsiloxane (PDMS) and fluorocarbon (FC) film layers. We demonstrated the successful pneumatic injection of a red ink solution into the separation channel as a proof of the concept. A mixture of fluorescein and dichlorofluorescein (DCF) could be baseline-separated using a single power source in microchip electrophoresis.

Electrophoresis, Microchip↗

PubMed2005

Microaffinity purification of proteins based on photolytic elution: toward an efficient microbead affinity chromatography on a chip.

A bead affinity chromatography system, which was based on the photolytic elution method, was integrated into a glass-silicon microchip to purify specific target proteins. CutiCore beads, which were coupled with a photo-cleavable ligand, such as biotin and an RNA aptamer, were introduced into a filter chamber in the microchip. The protein mixture containing target protein labeled with fluorescein isothiocyanate (FITC) was then passed through the packed affinity beads in the microchamber by pressure-driven flow. During the process, the adsorbed protein on the bead was monitored by fluorescence. The concentrated target protein on the affinity bead was released by simple irradiation with UV light at a wavelength of 360 nm, and subsequently eluted with the phosphate buffer flow. The eluted target protein was quantitatively detected via the fluorescence intensity measurements at the downstream of the capillary connected to the outlet of the microchip. The microaffinity purification allowed for a successful method for the identification of specific target proteins from a protein mixture. In addition, the feasibility of this system for use as a diagnosis chip was demonstrated.

Biotin↗

PubMed2004

On-line sample cleanup and chiral separation of gemifloxacin in a urinary solution using chiral crown ether as a chiral selector in microchip electrophoresis.

In chiral capillary electrophoresis of primary amine enantiomers using (+)-18-crown-6-tetracarboxylic acid (18C6H4) as a chiral selector, the presence of alkaline metal ions in the sample solution as well as in the run buffer is undesirable due to their strong competitive binding with 18C6H4. A channel-coupled microchip electrophoresis device was designed to clean up alkaline metal ions from a sample matrix for the chiral analysis of amine. In the first channel, the metal ions in the sample were monitored by indirect detection using quinine as a chromophore and drained to the waste. In the second separation channel, gemifloxacin enantiomers, free of the alkaline metal ions, were successfully separated using only a small amount of the chiral selector (50 microM 18C6H4).

Anti-Bacterial Agents↗

PubMed2004

Microbead-based affinity chromatography chip using RNA aptamer modified with photocleavable linker.

A microbead-based affinity chromatography chip (micro-BACC) controlling hundreds of nanoliters of reaction volume was developed to separate and analyze hepatitis C virus (HCV) RNA polymerase protein by immobilization of an RNA aptamer on beads. A photocleavable linker was conjugated in between the beads and the aptamer to elute the bound RNA polymerase from the RNA aptamer in one step by UV irradiation, resulting in an efficient method to elute and identify the target molecule bound on RNA using a mass spectrometer. This linker showed a cleavage activity over 70% upon UV irradiation at 1050 mW/cm2 for more than 5 min. The photoelution method could prevent the target molecule from contaminations in affinity chromatography caused by elution solutions of high salt concentration, extreme pH and detergent, respectively. In this chip, sample reagents up to 800 nL could be metered quantitatively into the bead chamber using a nanoliter dispenser working, based on surface-guided flow control and pneumatic control by external air pressure on the chip. RNA polymerase eluted after UV irradiation was successfully analyzed by trypsin treatment without additional purification. As a result, using the aptamer, we could detect RNA polymerase from 800 nL hepatitis C patient serum containing 96 fmol HCV RNA polymerase. The detection limit of this system was estimated to be 9.6 fmol HCV RNA polymerase.

Chromatography, Affinity↗

PubMed2003

Protein patterning by maskless photolithography on hydrophilic polymer-grafted surface.

With the help of a microfabrication process and surface modification technology, a method of fabricating protein patterned chips was developed which can be utilized as a powerful tool for performing bioassays in a high-throughput manner. A digital micromirror array (MMA) system was used as a virtual photomask, so that a maskless photolithography process was able to be used to build patterned biomolecules on a chip by selective illumination onto the chip surface. We utilized the nitroveratryloxycarbonyl (NVOC) group as a photolabile protecting group for protein patterning. The NVOC-protected surface was selectively irradiated by a UV illuminator using an MMA. After removing the NVOC group, biotin was coupled to the NVOC-cleaved site, onto which a buffered streptavidin solution was eluted. At this point, we could obtain a streptavidin-patterned surface and observe the effect of the polymer-grafted surface in reducing nonspecific binding.

Adsorption↗

PubMed2003

Integration of on-column immobilized enzyme reactor in microchip electrophoresis.

A simple method integrating an immobilized enzyme reactor into a microchip electrophoresis device was developed. The enzyme immobilization into a microchip was performed by spotting and drying a drop of dissolved nitrocellulose (NC) on a glass substrate, and adsorbing enzyme on the reconstituted NC membrane. This enzyme-immobilized glass plate was assembled with a polydimethylsiloxane substrate on which the separation channel was fabricated. The advantage of this method is the ability to easily change the position and size of the reactor within the microchip electrophoresis device. A beta-galactosidase reaction was demonstrated with fluorescein di-beta-D-galactopyranoside using this integrated on-column enzyme reactor. A successful electrophoretic separation of its hydrolysis products, i.e., fluorescein mono-beta-D-galactopyranoside (FMG) and fluorescein, was achieved. Enzyme kinetics and inhibition of the beta-galactosidase using FMG and 2-phenylethyl beta-D-thiogalactoside, respectively, were also studied with microchip electrophoresis.

Collodion↗

PubMed2003

Protein patterning on silicon-based surface using background hydrophobic thin film.

A new and convenient protein patterning method on silicon-based surface was developed for protein array by spin coating of hydrophobic thin film (CYTOP). Photolithographic lift-off process was used to display two-dimensional patterns of spatially hydrophilic region. The background hydrophobic thin film was used to suppress nonspecific protein binding, and the hydrophilic target protein binding region was chemically modified to introduce aldehyde group after removal of the photoresist layer. The difference in surface energy between the hydrophilic pattern and background hydrophobic film would induce easier covalent binding of proteins onto defined hydrophilic areas having physical and chemical constraints. Below 1 microg/ml of total protein concentration, the CYTOP hydrophobic film effectively suppressed nonspecific binding of the protein. During the process of protein patterning, inherent property of the hydrophobic thin film was not changed judging from static and dynamic contact angle survey. Quantitative analysis of the protein binding was demonstrated by streptavidin-biotin system.

Biosensing Techniques↗

PubMed2002

Chiral separation of gemifloxacin in sodium-containing media using chiral crown ether as a chiral selector by capillary and microchip electrophoresis.

Chiral crown ether, (+)-(18-crown-6)-tetracarboxylic acid (18C6H(4)), is an effective chiral selector for resolving enantiomeric primary amines owing to the difference in affinities between 18C6H(4) and each of the amine enantiomers. In addition to the destacking effect of sodium ion in the sample solution, the strong affinity of sodium ion to the polyether ring of crown ether is unfavorable to chiral capillary electrophoresis using 18C6H(4) as a chiral selector. In this report, the chiral separation of gemifloxacin dissolved in a saline sample matrix using 18C6H(4) was investigated. Adding a chelating agent, ethylenediaminetetraacetic acid (EDTA), to the run buffer greatly improved the separation efficiencies and peak shapes. The successful chiral separation of gemifloxacin in a urinary solution was demonstrated for both capillary and microchip electrophoresis.

Anti-Infective Agents↗

PubMedDate not supplied

Mechanical characterization of micro/nanoscale structures for MEMS/NEMS applications using nanoindentation techniques.

Mechanical properties of micro/nanoscale structures are needed to design reliable micro/nanoelectromechanical systems (MEMS/NEMS). Micro/nanomechanical characterization of bulk materials of undoped single-crystal silicon and thin films of undoped polysilicon, SiO(2), SiC, Ni-P, and Au have been carried out. Hardness, elastic modulus and scratch resistance of these materials were measured by nanoindentation and microscratching using a nanoindenter. Fracture toughness was measured by indentation using a Vickers indenter. Bending tests were performed on the nanoscale silicon beams, microscale Ni-P and Au beams using a depth-sensing nanoindenter. It is found that the SiC film exhibits higher hardness, elastic modulus and scratch resistance as compared to other materials. In the bending tests, the nanoscale Si beams failed in a brittle manner with a flat fracture surface. The notched Ni-P beam showed linear deformation behavior followed by abrupt failure. The Au beam showed elastic-plastic deformation behavior. FEM simulation can well predict the stress distribution in the beams studied. The nanoindentation, scratch and bending tests used in this study can be satisfactorily used to evaluate the mechanical properties of micro/nanoscale structures for use in MEMS/NEMS.

Journal Article↗