Earticle

The highly sensitive detection of pathogen in the soil samples is great importance because food poisoning outbreak has been occurred mostly from the pathogen infection of the soil. To detect low concentration of pathogen in a soil using PCR amplification, the removal process of humic substances is required because they are known as an inhibitor to PCR amplification. Various methods to remove humic substances in soil samples for nucleic acid based detection of pathogen have been developed. However, these methods don’ suitable the micro total analytical system. In this study, we’e developed the simple separation and concentration method of pathogen from the soil samples including humic acid for the nucleic acid based detection of pathogen. The humic acid was removed from the soil samples by optimal solution chemistry and physical adsorption. As a result, we could detect pathogen in low concentration in the soil sample included humic acid [<102 c.f.u/soil 10g]. Acknowledgment: This subject is supported by Ministry of Environment of the Republic of Korea as “he Eco-Technopia 21 project”ò and by the Korea Science and Engineering Foundation (KOSEF) grant funded by the Korea government (MEST) (No. 20100020657)).

A general method for the facile development of new biosensors for detection of liver biomarker protein targets have constructed and characterized. The peptides as recognition element selected using M13 phage display. A quartz crystal microbalance (QCM) is used as a diagnosis tool during biosensor development, and electrochemical techniques (cyclic voltammetry (CV) and electrochemical impedance spectroscopy (EIS)) are also used as the detection methods in the biosensor. A novel peptide capable of binding to alanine transaminase (ALT) was discovered by biopanning an M13 phage-displayed library. The selection procedure readily converged to a peptide (ALT5-8), with an amino acid sequence WHWRNPDFWYLK. The binding affinity of the peptide-containing phage particles was assessed by ELISA and the specific recognition of ALT was observed with a picomolar binding constant (Kapp= 8.14 pM). Peptides with the same sequence were chemically synthesized with a C-terminal cysteine and immobilized on a gold surface. These results demonstrate a simple platform for developing sensitive peptide-based biosensors for almost any desired protein targets.

The highly sensitive detection of norovirus and pathogenic bacteria in the food samples is of great importance because food poisoning outbreak has been wide occurred worldwide. For the molecular detection of a pathogen in the food samples, sample preparation procedures are required. Various methods to isolate a pathogen from the real samples have been reported. These methods, however, don’ fit for the micro total analytical system. Here, we’e developed the simple concentration method of norovirus from the oyster sample for the nucleic acid based detection of norovirus. The adsorption of norovirus on the solid surface was investigated with additives (ions, surfactants, pH) and surface characters (hydrophobicity and charge). As a result, we could isolate and concentrate norovirus from oyster samples (contains E.coli and protein), which results in that high sensitive detection of norovirus in low concentration in the oyster sample [101 PFU/a piece, PCR-calculated concentration] was achieved. Acknowledgment: This subject is supported by Ministry of Environment of the Republic of Korea as “he Eco-Technopia 21 project”and by the Korea Science and Engineering Foundation (KOSEF) grant funded by the Korea government (MEST) (No. 20100020657)).

In this work, we suggested the realization of supramolecular assembly of single walled carbon nanotubes (SWNTs) with the peptides for the directed growth and assembly of Au nanoparticles onto one-dimensional (1D) arrays. The programmed peptides containing phenylalanine and cystein groups can serve as bifunctions to create highly definable supramolecules of SWNTs and peptides and to guide the 1D arragement of Au NPs. We find that the highly stable dispersion and electrical properties of this conjugation were applied them into producing transparent conductive flexible thin films under vacuum filtration. As-obtained thin films were used as biosensing platforms for detecting biomolecules through electrochemical sensing method. The biosensor device shows a high sensitivity, low detection limit, and fast response time.

The recombinant bioluminescent bacteria which contained different promoters fused with either vibrio fisheri lux or bacterial thermorhabidus lux genes can emit bioluminescence upon the transcription initation at its proper temperatures either 30℃ or 37℃. We have evaluated total 29 different recombinant bioluminescent bacteria with gamma-ray irradiation in order to study their specific responses which act upon different stresses such as DNA-damage, oxidative damage, cell-wall damage, hydroxyl-radical damage, benzoic acid damage, and so on. Using 96 well-plates with different gamma-ray doses, we found that some bacteria containing DNA-damage sensitive promoters such as recA, SbmC, SulA, RecN have shown the sensitive bioluminescent from above the specific gamma-ray dose, while other DNA-damage sensitive bacteria containing NrdA or AlkA and other stress-response bacteria have not shown significant responses in their bioluminescence. It is expected that these distinct responses to gamma-ray irradiation can be used as biosensors for environment or space applications.

The promoter activity is a key factor in a cell for the active growth of living things and mass production of useful materials. To investigate the promoter activity of eukaryotic cell, in-vivo transcription has been usually used. However, for the faster monitoring of genetic information processing, in-vitro transcription process is essentially needed, because eukaryoticcell grows up slowly. Here we compared the several promoters activities by a simplified microalgal in-vitro transcription. RNA polymerase and the other cofactors were seperated from the chlamydomonas's cell lysate using MNP-DNA conjugates. Subsequently, concentration of mRNA produced by in-vitro transcription was measured by Qubit fluorometer. As a results, we could observe the changes in amounts of mRNA depends on the kinds of promoters. Thus, the best promoter could be selected. This system allows the fast,easy and economical analysis of promoter activity of microalgal under in-vitro transcription condition.

DNAzyme (also called deoxyribozyme or catalytic DNA) is a singlestranded deoxyribonucleic acid (DNA) that contains specific catalytic activity. Various DNAzymes are used in biosensing system to detect target nucleic acid or small molecule.1 We have developed a novel target nucleic acids detection method using Cu2+-dependent DNAzyme which cleaves single-stranded DNA in the presence of copper ion.2 The triplex region of DNAzyme was modified to contain target complementary sequences. The region was separated into two parts and prolonged with sequence which can bind to target. When target nucleic acid bound to modified DNAzyme, it formed stable structure and is activated to cleave substrate DNA. However, the DNAzyme cannot form active structure without target nucleic acid. For sensitive fluorescent detection using our system, the substrate DNA for DNAzyme was labeled with a FAM fluorophore at the 5' end and a quencher at the 3' end. Quenched FAM emission of substrate DNA increased by cleavage and release of FAM-labeled substrate from active DNAzyme with target. Single-stranded artificial target was successfully detected by our system at room temperature with high selectivity. Based on our study, we believe that this simple and cost-effective detection platform can be applied to various targets detection by only change in sequence of detection probe part, while fluorophore-labeled substrate DNA is universal.

The development of rapid and accurate bioassay methods for pathogenic bacteria detection is of great significance in all areas related to health and safety such as clinic, food industry, environment, and defense. As surface-enhanced Raman scattering (SERS) is a convenient and sensitive method that can detect the analytes even on a single molecule level using a simple nanostructure, it has been employing as an attractive strategy for sensitive DNA sensing. Here, we developed a SERS-based Au particle-on-wire system for multiplex identification of pathogenic bacteria1. The Au NW-on-wire exhibits superb reproducibility, excellent sensitivity, and good time stability. A pattern formed by multiplex Au NW sensors provides positional address and identification for each sensor. By using this system, reference strains and clinical isolates were successfully identified, predicting practical values of this sensor in a variety of biosensing application where DNA binding involved. [This research was supported by World Class University Program (R32- 2008-000-10142-0) through the Korea Science and Engineering Foundation funded by the Ministry of Education, Science and Technology]

Stable contacts between biomolecules and nano-materials are strongly required to design and characterize the bio-nano interface for many applications, such as medical implants, drug delivery, and biosensors. Although there are various strategies to construct the bio-nano interface to fabricate advanced devices with long lifetime, most of strategies are difficult to control the biomolecular binding and maintain the biological activity onto surface of nano-materials. Herein, we demonstrated the powerful and effective approach to attach organophosphrous hydrolase (OPH) onto graphene-based nanohybrids (RGO-Au) through site-specific interaction. The gold binding peptide (GBP)-OPH/RGO-Au was characterized by scanning electron microscopy and circular dichroism (CD). The secondary structure of enzymes after attachment onto RGO-Au represented negligible changes with the GBP. In addition, the GBP-OPH/RGO-Au hybrid were applied into biosensors for detecting organophosphorous compounds.

Quantification of mRNA provides information crucial for various biological studies. Real-time PCR is known to be the most accurate method for quantifying mRNA, and thus represents the state-of-the-art for gene expression analysis. However, the use of real-time PCR for mRNA quantification is limited to a single target per analytical run because of reductions in quantification power and limitations of fluorescence dyes associated with multiplex applications. Capillary electrophoresis-single-strand conformation polymorphism analysis (CE-SSCP) is an alternative multiplex analysis method. However, CE-SSCP has not been widely used for multiplex applications due to low resolution problem. In this study, we developed high-resolution CE-SSCP system using PEO-PPO-PEO triblock copolymer solution. Moreover, for the multiplex amplification of RNA, modified multiplex ligation-dependent probe amplification (MLPA) was combined with CE-SSCP analysis so that the amount of mRNA could be quantified precisely. We have demonstrated that MLPA-CE-SSCP could be used to monitor expression of 31 metabolic genes of Escherichia coli, 16 genes of Caenorhabditis elegans and 22 genes of Arabidopsis thaliana.

The combination of Localized Surface Plasmon Resonance (LSPR) with interferometry using gold-capped porous anodic alumina (PAA) layer chip for the detection of C-reactive protein (CRP) antibody-antigen interactions is demonstrated. The combination of LSPR with interferometry on a gold capped PAA layer chip enables two important aspects of optical sensing systems, the shift in λmax and the increment in the absorbance intensity in a new and highly sensitive format. The excitation of the optical characteristics and the detection was performed using only one optical fiber, which made our device user-friendly and suitable for the development of hand-held diagnostic devices. For the fabrication of PAA layer chip, we developed a two-step anodizing method. The porous portion of PAA layer chip consisted of close-packed hexagonal inter-pores, each with a central straight nano-pore perpendicular to the surface of Al surface. Diameters of the nano-pore and the cell were about 60 nm (1.2X1010pores/cm2) and 120 nm, respectively. Then, PAA layer chip surface was deposited with gold by using thermal evaporator. The CRP antibody immobilization was carried out with the functional SAM on the surface of gold-capped PAA layer chip. For the measurement of CRP antibody-antigen interactions, small volumes of label-free CRP antigen were applied onto the surface of gold-capped PAA layer chip. Subsequently, evaluation of optical properties was carried out. In the presence of CRP antigen, the change of absorbance intensity in the interferometric pattern and a significant bathochromic shift was observed.

Polydiacetylene (PDA) liposomes which possess unique properties and that properties allow liposomes to change color and emit fluorescence in response to stimulus such as temperature, antibody-antigen interaction, variations in pH, mechanical stress and organic solvent. This material has been extensively studied as signal transducers in biosensor applications. In this research, we demonstrated an antibody-based biosensor using PDA liposomes for detection of hIgE. Target hIgE chemically bound to antibodies-immobilized PDA liposomes and the fluorescence responses were a little bit increased depending on target protein concentration. On the other hand, in case of the addition of gold nano particles-conjugated polyclonal antibody on the liposomes, fluorescent responses dramatically increased depending on target protein concentration. As the results, we could confirm that gold nano particles-conjugated polyclonal antibody affect fluorescent signal amplification of PDA liposomes chip. This strategy can be useful to detect proteins of ultra low concentration.

Rheumatoid arthritis (RA) is an autoimmune disease by persistent synovitis, systemic inflammation, and autoantibodies that may affect many tissues and organs, but principally attacks synovial joints. RA is a chronic illness, patients may experience long periods without symptoms however it causes joint destruction and functional disability. RA is difficult to diagnose in its early stages for several reasons. First, RA difficulties to distinguish established RA from other types of established joint diseases. Second, there is no single test for the disease. Therefore, early diagnosis of RA is essential for early treatment of RA. We were designed diagnosis system using RA biomarker and aptamer. Previous studies reported an increase of tumour necrosis factor (TNF) protein when RA occurred in patients. Overproduction of TNF leads to overproduction of many cytokines such as interleukin 6, which also drives persistent inflammation and joint destruction. Therefore, we cloned the RA biomarker candidate gene (TNF) and obtained TNF target protein by heterologous expression system. The TNF specific aptamer selected to develop of RA early diagnosis system. In conclusion, the developments of TNF detection system using specific binding aptamer for RA biomarker can be applied to early diagnosis and early pertinent treatment of RA.

Synthesis of intelligent genetic network in bacteria is one of the great applications of synthetic biology. Here we show the assembly of an intelligent system that can trigger bacteria to sense and respond to the removal of heavy metals such as zinc and copper by engineering of two component systems. This bacterial system can sense zinc and copper using OmpR and NtrC family of two component sensor system and remove metals contaminants through cell surface display system. The genetic circuit of HydHG and CusSR were fused to cell surface display system for the expression of chimeric protein namely OmpC with heavy metal binding peptide. Our Intelligent bacterial system targets the specific heavy metal and triggers a cell surface display system in response to the heavy metal added to the bacterial population which is potentially used in the field of biosensor and bioremediation.

An immunoassay in which mycotoxin ochratoxin A (OTA) coupled with anti-OTA antibodies for generating fluorescence resonance energy transfer (FRET) was developed, optimized, and validated. At neutral pH or lower pH condition, FRET was observed between the OTA and anti-OTA, because of their intrinsic fluorescence properties. Based on the unique fluorescence properties, the OTA was able to detect 1 ng/ml with high sensitivity and specificity. Furthermore, a weak cross-reaction for ochratoxin B (OTB) was observed. In addition, the assay required only within 30 min to obtain results and one step to perform the assay. In validation, the results obtained from spiked coffee and wheat samples by the FRET immunoassay were in good agreement with those obtained by direct competitive enzyme-linked immunosorbent assay. Therefore, the results obtained in this study could be used as basic research for the development of FRET immunoassay and as a useful on-site screening tool for the rapid detection of OTA in wheat, coffee, and dried fruits.

Green Fluorescent Protein (GFP) is an indispensable tool with a wide range of application as reporter protein in the cellular and biological studies. It helps in direct visualization of bimolecular and biochemical activities of the living cells. Although a wide range of GFP variant has been generated, developing a far red fluorescent protein in an indispensable quest in the cell biology for the purpose of deep tissue and whole cell imaging. For this purpose, enormous effort has been done by the researchers to generate far red fluorescent proteins through incorporation of natural and unnatural amino acid in the chromophore of GFP. In this study, we have generated a highly red shifted GFP by incorporating the tyrosine analog, amino tyrosine which provides the GFP with the inherent characteristics of Red fluorescent protein (RFP) such as emission maximum at longer wavelength and copper sensing characteristics. This study offers an insight into the spectral properties of amino tyrosine incorporated GFP and the mechanism of fluorescence quenching in response to copper concentration. The Copper binding property of the engineered protein was characterized by the combination of fluorescent spectroscopy, absorption spectroscopy and circular dichroism. Overall this study demonstrates that the development of red shifted GFP with inherent characteristics of red fluorescent protein.

The human immunodeficiency virus (HIV) pandemic mainly affects developing countries, where the Joint United Nations Programme on HIV/AIDS (UNAIDS) estimates suggest that less than 1 in 10 people are aware of their HIV sero-status. In order to enhance epidemiological surveys, prevention programs, and therapeutic interventions, development of specific, rapid, and convenient diagnostic detection systems for HIV is still warranted. Here we report the direct detection of HIV particles using broadly HIV-1 neutralizing gp120 antibody (2G12, gp120MAbs)-conjugated magnetic beads (MBs) and fluorescent nanosized polymeric beads (FNBs). As a model target, noninfectious pseudo HIV particles (pHIVPs) were produced through cotranspected HEK293 by the two plasmids, pCMV-dR.8.74 and pDOLHIVenv, which contain Gag and Pol, and Env of HIV-1, respectively. In the presence of pHIVPs, addition of gp120MAbs-FNBs and gp120MAbs-MBs leads to the formation of sandwich complex, which can be easily isolated and concentrated by common magnet separation. We demonstrate the ability of detecting HIV-1 particles specifically and directly with low sample volume (less than 100 μL) and rapidity (less than 1.5 h). These results suggest that broadly HIV-1 neutralizing antibodies coupled to nanobeads can be employed for the direct detection of HIV-1 particles with potential implication for the development of specific, rapid, and convenient diagnostic systems.

Artificial taste sensors, so called ‘lectronic tongues’ have been widely used in food and beverage industry. The electronic tongues usually utilize synthetic materials for the recognition of analytes. In this reason, they cannot mimic the biological features of the human taste system. To overcome these limitations, we applied biological human taste receptor for the development of artificial taste sensor. Here, we demonstrated human bitter taste receptor, hTAS2R38, functionalized conducting polymer nanotube(CPNT)-field effect transistor (FET) mimicking human taste system. Each taster (PAV) and non-taster (AVI) haprotype of hTAS2R38 genes were cloned into bacterial expression vector and then expressed in E. coli at high-level. The receptors were then immobilized on CPNT-FET sensor platform. The PAV-CPNT-FET responded to target bitterness compounds, phenylthiocarbamide (PTC) and propylthiouracil (PROP), with high sensitivity down to concentration as low as 10 fM. However, AVI-CPNT-FET did not stimulated by target bitter tastants. This artificial taste sensor showed very similar performance with human taste system and may offer a powerful tool for basic research on taste receptors.

We introduced Flow through hole(FTH) system that can be measured a wide range of concentrations of the antigen and relatively small volume of samples with human serum within minutes. This system is designed to the vertical direction, instead of existing horizontal direction like lateral flow assay (LFA). As immunoassay reaction based on gold nanoparticles(AuNP), this reaction occurs while the sample solution passes through hundreds of micro-size holes. Under optimal conditions, FTH system can rapidly detect C-reactive Protein(CRP) antigen from 10 ng/mL to 10 μg/mL. It is observed that signal intensity increased until 2min and then equally was maintained from 2 min to 10 min by saturation of AuNP complex. That is, immunoassay reaction is completed within 2 min and then any additional sample flow does not effected signal intensity change. As compared to the conventional LFA, the detection time sucessfully shortened.

Single-walled carbon nanotubes (swCNTs) hold great promise for use as molecular wires because they exhibit high electrical conductivity and chemical stability. However, constructing swCNT-based transducer devices requires controlled strategies for assembling biomolecules on swCNTs. In this study, we developed a chemically modified swCNT for reliable attachment of the human olfactory receptor (hOR) onto the CNTs and investigated their use in the fabrication of a highly sensitive and selective bioelectronic nose. For the bioelectronic nose, the swCNT-field effect transistor platform was composed of polyethylene glycol-coated regions to prevent non-specific absorption and chemically modified swCNTs regions containing hOR, which can bind to the specific odorant. This approach allowed us to create well-defined micron-scale patterns of hOR on the swCNTs. Our bioelectronic nose displayed ultrahigh sensitivity down to concentrations as low as 1 fM. In addition, the approach described here may provide an alternative route for multiplexed detection of diverse odorants and to improve the sensitivity of sensor devices.

Herein, a new electrochemical method for the analysis of methyltransferase activity was developed based on enzymatic methylation/cleavage coupling reaction1,2 utilizing quantum dot as a signaling tracer3. On the gold matrix immobilized with ssDNA, biotin-modified complementary strand and streptavidin-coated CdSe/ZnS QDs were successively reacted resulting in QD tethered dsDNA. Then, various concentration of Dam (DNA adenine methylation) MTase was applied which generated methylated sequence(GAmTC) in dsDNA. For the signaling, DpnI was applied to the methylated dsDNA to specifically cleave the methylated site generating free QD which could be detected by anodic stripping voltammetry. As increasing amount of methylated DNA, electrochemical signal generated from QD increase because the amount of free QDs increase. As a result, MTase activity can be simply detected with high sensitivity, selectivity and wide linear range(8-160U/mL). This is the first study of the electrochemical method for MTase activity analysis, furthermore, our strategy could be used as a basic key technology in the field of epigenetics.

We developed a multi-bit biomemory chip which was consisted of recombinant azurin variants. To store the different information for memory performance, this recombinant azurin was substituted from Cu ion to various metal ions. Each metal-substituted recombinant protein is directly self-assembled onto Au surface via thiol group of thecysteine by covalent bonding. A cyclic voltammetry (CV) and an open circuit potential (OCP) indicated that recombinant azurin variants have different redox peaks and specific open circuit potential values. Using these parameters, the memory function was verified by a chronoamperometry (CA) and open circuit potential amperomemtry (OCPA). These results presented here provide a new dimension, concept and material combinantion for developing new-type memory systems with recombinant protein. Acknowledgments : This research was supported by The Nano/Bio Science &Technology Program (M10536090001-05N3609-00110) of the Ministry of Education, Science and Technology (MEST), by the Original Technology Research Program for Brain Science through the National Research Foundation of Korea (NRF) funded by the Ministry of Education, Science and Technology (2009-0093907), by the Ministry of Knowledge Economy (MKE) and Korea Institute for Advancement in Technology (KIAT) through the Workforce Development Program in Strategic Technology.

A biosensor based on the localized surface plasmon resonance (LSPR) response of a single Au nanoparticle was fabricated for the highly sensitive detection and quantification of a specific cancer biomarker. The spectral position changes of single Au nanoparticles induced by the interaction of adsorbates and target analytes were effectively utilized as sensing tools. The LSPR responses of single Au nanoparticles were obtained by tracking the wavelength shift of the corresponding resonant Rayleigh-light-scattering spectra via dark-field microspectroscopy. Using prostate specific antigen as a model, an LSPR λmax shift of about 2.75 nm was recorded by a primary immunoresponse corresponding to 0.1 pg/mL of the target antigen. The sensitivity of the immunoassay can be substantially enhanced, however, by a sandwich strategy. A PSA polyclonal antibody was used as an amplifying agent in the strategy. As a result, the linear dynamic range of the sensing platform was determined to be within the concentration range of 10-4 to 0.1 ng/mL and a detectable minimum concentration of 0.1 pg/mL was identified, with an LSPR λmax shift of about 4.96 nm. The results indicate that the aforementioned approach can significantly contribute to the fabrication of ultrasensitive biosensors, allowing the quantitative analysis of cancerassociated proteins.

We report on the development of an efficient optical-sensing platform for the detection of toxic organophosphates using self-assembled peptide nanotubes. Diphenylalanine (Phe-Phe, FF), found in the key motif of Alzheimer’ b-amyloid, was used as a building block to create self-assembled nanotubes that became highly photoluminescent through in-situ incorporation of lanthanide complexes such as salicylic acid/terbium ions or 1,10- phenanthroline/europium ions during the self-assembly process. We found that photoluminescent FF nanotubes underwent a drastic quenching of their emission within a few seconds upon exposure to paraoxon, a nitrofunctionalized neurotoxin. The photoluminescence quenching occurred due to the interruption of cascaded-energy transfer from FF nanotubes to lanthanide ions. The assay platform provided high selectivity toward paraoxon among other organophosphates (e.g., diethylchlorophosphate, dichlorvos, malathion, phosmat), nitro-group compounds (e.g., nitromethane, nitrobenzene, 4-nitrophenol) and common organic chemicals (e.g., alcohols, ethers, ketones, etc.), capability of which was attributed to the role of the FF nanotubes as a host matrix for lanthanide complexes. The peptide nanotube-based optical sensor hints at an alternative route for the selective detection of neurotoxins while avoiding low selectivity, slow response time, and instability of biological receptors in other biosensors.

The abnormal aggregation of β-amyloid peptide (Aβ) in the brain is a major histopathological feature of Alzheimer’ disease (AD). Herein, we report on microfluidic dissociation and clearance analysis of preformed Aβ aggregates for parallel screening of aggregate destabilizers in a high-throughput manner. As a proof of the concept for the microfluidic platform, we investigated (1) microfluidics-based clearance of metal ion-induced Aβ aggregates using different types of metal chelators, (2) the clearance effect of deferoxamine on Aβ aggregates within microchannels, (3) comparison between destabilized Aβ dissociated from preformed A β aggregates and remaining deposits within the microchannels both before and after the clearance, and (4) secondary structure change in Aβ deposits by the clearance treatment. The microfluidics-based clearance system should be suitable for high-throughput screening of drug candidates to enhance the clearance of Aβdeposits for AD therapy prior to their in vivo evaluation.

We constructed an HPLC-type system suitable for quantitative analysis that enables base sequence- and Tm-dependent DNA oligomer separation using a small open tubular capillary column (300 mm×100 μm I.D.). The DNA oligomers with an amino group at the 5'-end (used as probes) were immobilized on the inner silica surface of the capillary column which had been sequentially treated with APTS (3-aminopropyltriethoxysilane), BMS (butyltrimethoxysilane), and DSG (disuccinimidylglutarate). We succeeded in separating complementary and non-complementary DNA oligomers in specific and quantitative modes. We also designed a temperature gradient strategy for efficient separation of target DNA oligomers. Using a column carrying two different probes with similar Tm values, their complementary target DNA oligomers were also separated and detected. The developed DNA open tubular capillary column system developed in this study could be further improved and used for the quantitative analysis of DNA or mRNA samples.

Oxygen is the main reason for food spoilage not only because it can chemically react with food but also because its presence is vital for the growth of food spoilage organism. Hence, modified atmosphere packaging (MAP) is born to elongate food shelf-life via reducing the oxygen concentration in package. Along with MAP is the demand for easy and quick analytical methods for detecting the presence of oxygen. This oxygen indicator should be non-toxic, water insoluble, cheap, reliable and sensitive. A solvent-based, irreversible colorimetric ink - consisting of a photosensitiser SnO2, a mild reducing agent, a redox dye, and an encapsulating polymer - was described in this research. Based on this ink, an ink-coated film lost its color rapidly upon exposure to UVB and remains colorless in absence of O2, returning to its original color in the presence of oxygen. Unlike its TiO2 - based predecessor, this ink prevented the undesirable reactivation by UVA light from white fluorescent light in food cabinet. Effects of type and ratio of the ink components were examined to improve the reliability and sensitivity of the oxygen indicator. The m echanism of this indicator along with its potential application was also discussed.

Electric Cell-substrate Impedance Sensing (ECIS) has been currently emerging as a novel, non-invasively and sensitive techniques in detection and analyzation of cellular responses to chemical and biological agent. In this study, we provide the fabrication and measurement techniques of ECIS system to detect cell viability and cell-toxicity screening. A bi-layer of Au/Ti was first deposisted and etched to form the sensing area, lead trace and the contact pad. The sensing microelectrodes were then photolithographically defined by a thin dielectric layer of SU-8 photoresist. For carrying out the detection and measurement, the HeLa cells were cultured on these electrodes and alternative current (AC) with the frequency from 100Hz to 100KHz was applied. The cellular spreading, morphology and multiplication was monitored by the impedance changes of the obtained electrical signal. At last, the anti-cancer drug was applied to watch for the cell-toxicity screening. With this design of ECIS system, we are able to use the commonly used ECIS-based technique to integrate into the microfluidic devices for many various purposes. Acknowledgment: This subject is supported by the Korea Science and Engineering Foundation (KOSEF) grant funded by the Korea government (MEST) (No. 20100020657)

A electrical immunosensor based on sub-micro interdigitated electrodes (IDEs) and gold enhancement is developed for detection of human interleukin 5 (IL5). IDEs, facing comb-shape electrodes, can act as simple and miniaturized devices for immune sensing. An IDE with a gap size of 400 nm was fabricated by a stepper photolithography process due to its mass-producible and cost-effective advantages and was applied for the immunoassay of human IL5. A biotinylated anti-human IL5 was immobilized on the streptavidin-modified IDE, and biotin-bovine serum albumin (BSA) and BSA were added sequentially to reduce non-specific binding between the streptavidin-immobilized IDE surface and other proteins. The immunoassay procedure included three main steps: the reaction of human IL5 to form antigen-antibody complexes, the binding of AuNP conjugation with an antibody against human IL5 for the sandwich immunoassay, and gold enhancement for electrical signal amplification. The measurement of electrical current at each step showed that the gold enhancement step was very critical in detection of the concentration of human IL5. Analysis by scanning electron microscope (SEM) showed that close to 1 mm particles were formed from 10 nm AuNP by the gold enhancement reaction using gold ions and hydroxylamine. Under optimized conditions, human IL5 could be analyzed at 1 pg mL-1 with a wide dynamic range (from 10-3 to 100 ng mL-1 concentrations).

The immobilized enzyme is necessary for the development of enzyme fuel cells (EFCs) which can be applied to power source of implantable devices. The main issue to be solved is the low generation of electrical power by enzyme electrode due to several problems. One of them is the possible attachment of active site of enzyme onto support materials during immobilization of enzymes, which may decrease the chances to react between substrates and enzymes, thereby decreasing power density. Here, active site of enzyme (glucose oxidase) was masked by glucose or gluconic acid to protect active site from attachment on support material during immobilization on multiwall carbon nanotube. It was found that electric power density was increased by enzyme electrode prepared by this novel immobilization method.