• 센서학회지
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• 제20권1호
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• pp.35-39
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• 2011

A disposable electrochemical immunosensor system has been developed for the detection of herbicide in aqueous samples. Disposable screen printed carbon electrodes(SPCE) were used as basic electrodes and an enzyme, horseradish peroxidase (HRP), and anti-herbicide antibodies was immobilised on to the working electrode of SPCE by using avidin-biotin coupling reactions. An herbicide-glucose oxidase conjugates have been used for the competitive immunoreaction with sample herbicides. The enzymatic reaction between the conjugated glucose oxidase and glucose added generates hydrogen peroxide, which was reduced by the peroxidase immobilised. The latter process caused an electrical current change, due to direct re-reduction of peroxidase by a direct electron transfer mechanism, which was measured to determine the herbicides in the sample. The optimal operational condition was found to be: $20\;{\mu}gl-1$ deglycosylated avidin loading to the working electrode and working potential +50 mV vs. Ag/AgCl. The total assay time was 15 min after sample addition. The detection limits for herbicides, atrazine and simazine, were found to be 3 ppb and 10 ppb, respectively.

• 센서학회지
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• 제23권5호
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• pp.295-298
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• 2014

A novel approach to fabricating high-performance glucose sensors is reported, which is based on the process of self-assembled monolayers (SAMs). In this study, we have particularly used ${\alpha}$-lipoic acid (LA) SAMs for the glucose sensors. To our best knowledge, this study is the first one to use LA as SAMs for this purpose. N-hydroxysuccinimide (NHS) and 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide (EDC) were deliberately attached at the same time on the LA SAM. Then, glucose oxidase ($GO_X$) and horseradish peroxidase (HRP) were sequentially immobilized. Thus, the HRP/$GO_X$/NHS-EDC/LA-SAM/Au/Cr/glass working electrode was developed. The glucose-sensing capability of the fabricated sensor was systematically measured by the use of cyclic voltammetry in the range of 1-30 mM glucose in phosphate-buffered saline. The result showed a good sensitivity, that is, as high as $27.5{\mu}A/(mM{\cdot}cm^2)$. This result conspicuously demonstrates that LA can be one of promising substances for use as SAMs for accurately monitoring trace levels of glucose concentration in human blood.

• Journal of Microbiology and Biotechnology
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• 제14권3호
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• pp.612-619
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• 2004

A competitive direct enzyme-linked immunosorbent assay (cdELISA) was developed for selective and rapid detection of a glycopeptide antibiotic, teicoplanin (TP). TP was conjugated to bovine serum albumin (BSA) for use as an immunogen. Repeated subcutaneous injections of 0.5 mg of the conjugate was effective in generating specific polyclonal antibody (PAb) toward TP in rabbits, as determined by cdELISA. TP-horseradish peroxidase conjugate (TP-HRP) was used as an enzyme marker. The cdELISA was developed based on a competition reaction between TP-BSA PAb and TP-HRP conjugate. The TP-BSA PAb was highly sensitive (detection limit, 0.3 ng/ml and specific toward teicoplanin, showing no cross-reactivity to other glycopeptide antibiotics including vancomycin. There were good correlations ($r^2$=0.84 and 0.76, respectively) between cdELISA and microbiological assay, and high-performance liquid chromatography. The cdELISA system developed in this work is expected to be useful not only for selective and rapid monitoring of TP but also for study of TP pharmacokinetics.

• 전기화학회지
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• 제10권3호
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• pp.229-232
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• 2007

An amperometric immunosensor for the determination of rabbit IgG is proposed. The immunoassay utilizes a screen-printed carbon electrode on which osmium redox polymer is electrodeposited. This immunoassay detects 0.1 ng/ml of rabbit IgG, which is ${\sim}10^2$ fold higher than the most sensitive enzyme amplified amperometric immunoassay. The assay utilizes a screen-printed carbon electrode which was pre-coated by a co-electrodeposited film of an electron conducting redox hydrogel and a rabbit IgG. The rabbit IgG in the electron conducting film conjugates captures, when present, the anti-rabbit IgG. The captured anti-rabbit-IgG is labeled with horseradish peroxidase (HRP) which catalyzes the two-electron reduction of $H_2O_2$ to water. Because the redox hydrogel electrically connects HRP reaction centers to the electrode, completion of the sandwich converts the film from non-electrocatalytic to electro-catalytic for the reduction of $H_2O_2$ to $H_2O$ when the electrode is poised at 200 mV vs. Ag/AgCl.

• BMB Reports
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• 제29권3호
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• pp.192-199
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• 1996

A rapid and sensitive method has been developed to detect a-fetoprotein (AFP) in human serum by a two-site sandwich enzyme-linked immunosorbent assay (ELISA) with monoclonal antibodies (MAbs) for human AFP within 1 h. To obtain the most sensitive and reliable MAbs. 12 kinds of MAbs (HPJ1 to HPJ12) as a capture antibody and 4 kinds of horseradish peroxidase (HRP) conjugated MAbs as a tracer antibody were investigated. Among these, only HPJ 10-HRP conjugated HPJ 1 (HPJ 10-HPJ $1^*$) and HPJ 11-HRP conjugated HPJ 10 (HPJ 11-HPJ $10^*$) were chosen as candidates based on the linearity of the standard curve and the sensitivity of the assay. To further characterize these two pairs. MAbs against human AFP were purified from hybridoma cells. conjugated with HRP. and then characterized to optimize the two-site sandwich ELISA The HPJ 10-HPJ $1^*$ pair showed a sensitivity of 1 ng/ml and a better reproducibility than the HPJ 11-HPJ $10^*$ pair when the human sera were incubated at $37^{\circ}C$ for 30 min. The results obtained for 480 randomly selected human sera showed 0~20 ng/ml of AFP values for the normal human sera. To test the utility of our kit, AFP concentrations were determined for 951 human sera (including 85 normal sera, 480 random blood sera, 213 HBsAg-positives. 50 anti-HCV antibody positives. and 47 malignant diseases) and compared with other commercially available AFP detecting kits. These results show that the present two-site sandwich ELISA method is a rapid, sensitive, and reliable procedure for detecting AFP in human serum.

• Journal of Microbiology and Biotechnology
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• 제17권11호
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• pp.1826-1832
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• 2007

We describe the fabrication of poly(ethylene glycol) diacrylate (PEG-DA) hydrogel microstructures with a high aspect ratio and the use of hydrogel microstructures containing the enzyme ${\beta}$-galactosidase (${\beta}$-Gal) or glucose oxidase (GOx)/horseradish peroxidase (HRP) as biosensing components for the simultaneous detection of multiple analytes. The diameters of the hydrogel microstructures were almost the same at the top and at the bottom, indicating that no differential curing occurred through the thickness of the hydrogel microstructure. Using the hydrogel microstructures as microreactors, ${\beta}$-Gal or GOx/HRP was trapped in the hydrogel array, and the time-dependent fluorescence intensities of the hydrogel array were investigated to determine the dynamic uptake of substrates into the PEG-DA hydrogel. The time required to reach steady-state fluorescence by glucose diffusing into the hydrogel and its enzymatic reactions with GOx and HRP was half the time required for resorufin ${\beta}$-D-galactopyranoside (RGB) when used as the substrate for ${\beta}$-Gal. Spatially addressed hydrogel microarrays containing different enzymes were micropatterned for the simultaneous detection of multiple analytes, and glucose and RGB solutions were incubated as substrates. These results indicate that there was no cross-talk between the ${\beta}$-Gal-immobilizing hydrogel micropatches and the GOx/HRP-immobilizing micropatches.

• 한국미생물학회:학술대회논문집
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• 한국미생물학회 2001년도 추계학술대회
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• pp.121-127
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• 2001

Seven cyanobacteria strains (Anabaena macrospora NIERl0016, Oscillatoria sp. NIER10042, Microcystis aeruginosa NIER10015, M. ichtyoblabe BIER10025, BIER10040, M. novacekii NIER10029, M. wesenbergii NIER10068) were tested with four rRNA - targeted oligonucleotide probes labelled with horseradish peroxidase (HRP) and specific for cyanobacteria. Non- fluorescent detection of hybridization signal was used. The hybridization with artificial mixture of cyanobacteria have shown the possibility to use 2 species-specific probes in duplicate hybridization and detection with different colored substrates.

• 센서학회지
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• 제17권1호
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• pp.29-34
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• 2008

A new disposable amperometric tri-enzyme biosensor for the detection of paracetamol has been developed. The paracetamol sensors developed uses horseradish peroxidase modified screen-printed carbon electrodes (HRP-SPCEs) coupled with immobilized enzymes, tyrosinase and aryl acylamidase, prepared using a poly (vinyl alcohol) bearing styrylpyridinium groups (PVA-SbQ) matrix. Optimization of the experimental parameters has been performed and the paracetamol biosensor showed detection limit for paracetamol is as low as $100{\mu}M$ and the sensitivity of the sensor is $1.46nA{\mu}M^{-1}cm^{-2}$.

• 한국가축번식학회지
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• 제17권4호
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• pp.347-356
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• 1994

A simpled and sensitive microplate enzyme immunoassay(EIA) was developed for the determination of progesterone concentration in serum, based on progesterone monoclonal antibody as anti-progesterone, horseradish peroxidase(HRP) as enzyme-label and tetramethylbenzidine(TMB) as substrate. The assay has a sensitivity of 5 pg-120pg/well and intra- and inter-assay coefficients of variation for progesterone standard curve (1.0ng~10.0ng/ml) were ranged 2.5~9.9% and 1.7.8.0%, respectively, determination coefficient of the regressio equation of our standard curve(R2=0.990$\pm$0.007) were high, and this is the same level as that of commercial kit(Hormonost Bio-Lab, Germany, R2=0.98~0.99). The progesterone concentration of serum determined by both kits (Work & Bio-Lab) were significantly correlated (r=0.95, P<0.01) although a little higher value were resulted in our kit than that of commercial kit. It generally is these results indicated that the microplate-EIA can be cused for the determination of progesterone in serum, as well as, for the determination of the early pregnancy diagnosis.

• 한국식품위생안전성학회지
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• 제10권4호
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• pp.213-217
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• 1995

A screening method has been developed for detecting sulfamethazine(SMZ) contamination of meat or feeds by using horseradish peroxidase (HRP) labeled protein A (Prot AHRP)and an indirect competitve enzyme-linked immunosorbent assay(ELISA). The assay is based on competitve binding of guinea pig anti-SMZ with SMZ in smaple and SMZ-gelatin conjugate(SMZ.GEL). Percent binding (B.Bo$\times$100) was calculated from the absorbance in the absence (B0) and presence (B) of SMZ. By the sandard curve prepared by plotting log(SMZ) vs percent binding of each known reference solution, the detection limit was 1.0ppb or less. Cross reacton with sulfadimethoxine, sulfaguaniding, sulfamerazine, sulfamthoxpyridazine, sulfanilamide, sulfisomidine and sufisoxazole were not observed. But sulfamerazine crossreacted in the test. The EC-50 value (concentration causing 50% inhibition of color development compared with blank) of sulfamerazine was 2.0 ppm. Further quality control will make the ELISA system ideal for the detection of SMZ in meat or feeds.