• Journal of Sensor Science and Technology
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• v.20 no.6
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• pp.400-405
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• 2011

This study introduced the development of a strip type disposable enzyme-linked immunosensor platform for the detection of IgG. Strips of the strip sensor were fabricated by using commercial nitrocellulose filter membranes and a housing holder for the strips was manufactured by using a standard injection molding process for a plastic material. An IgG-urease conjugate was prepared and used for the competitive immune-binding with sample IgG. From the enzymatic reaction between the conjugated urease and urea added, ammonia was generated and caused a localized alkaline pH change on the immobilized antibody band which was coated onto the sensor strips. This pH increase subsequently caused a color change of the antibody band in the presence of a pH indicator, phenol red. Used in conjunction with a competitive immunoassay format, the intensity of the color produced is directly linked with the concentration of target analyte, IgG, and specific measurement of IgG in a lateral flow immunoassay format was achieved over the range 100 ppb to 2000 ppb IgG.

• Journal of Microbiology and Biotechnology
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• v.17 no.9
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• pp.1445-1451
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• 2007

A sensitive optical waveguide lightmode spectroscopy-based immunosensor was developed to detect vitellogenin in seawater flatfish (Paralichthys olivaceus). For this purpose, anion-exchange column chromatography with DE-52 resin was used to purify flatfish vitellogenin from flatfish serum containing vitellogenin that had been induced using an intraperitoneal $17{\beta}$-estradiol injection. The anti-flatfish vitellogenin antibody used as the biological component of the above immunosensor was prepared using the purified flatfish vitellogenin. The change in the incoupling angle according to the complexation between the flatfish vitellogenin and its antibody, immobilized over an optical grating coupler sensor chip, was measured to calculate the sensor response. The immunosensor was quite specific to flatfish vitellogenin binding, based on no sensor response in the case of bovine serum albumin immobilization. When plotted using double-logarithmic scales, the sensor responses increased linearly in flatfish vitellogenin concentrations of 0.00675-67.5 nM, with a detection limit of 0.0675 nM. The reusability during seven repetitive measurements was reasonably fair for the preliminary screening of flatfish vitellogenin.

• Journal of Biosystems Engineering
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• v.34 no.4
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• pp.254-259
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• 2009

Conventional methods for pathogen detection and identification are labor-intensive and take several days to complete. Recently developed biosensors have shown potential for the rapid detection of foodborne pathogens. In this study, an impedimetric biosensor was developed for rapid detection of Salmonella typhimurium. To develop the biosensor, an interdigitated microelectrode (IME) was fabricated by using semiconductor fabrication process. Anti-Salmonella antibodies were immobilized based on either avidin-biotin binding or self assembled monolayer (SAM) on the surface of the IME to form an active sensing layer. To evaluate effect of antibody immobilization methods on sensitivity of the sensor, detection limit of the biosensor was analyzed with Salmonella samples innoculated in phosphate buffered saline (PBS) or food extract. The impedimetric biosensor based on SAM immobilization method produced better detection limit. The biosensor could detect 107 CFU/mL of Salmonella in pork meat extract. This method may provide a simple, rapid, and sensitive method to detect foodborne pathogens.

• The Transactions of The Korean Institute of Electrical Engineers
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• v.65 no.4
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• pp.628-634
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• 2016

This paper demonstrates a microfluidic electrochemical sensor for detecting endocrine disruptor such as estradiol at a very low concentration by using preconcentration technique. In addition, self-assembled monolayer(SAM) was also employed on the working electrode of the electrochemical sensor in order to increase the estradiol capture efficiency of the sensor. SAM treatment on the working electrode enhanced the specific binding between the surface of the working electrode and the estradiol antibody. The estradiol antibody was applied on the working electrode at different concentrations(10, 20, 50, 100, 200 pg/ml) for observing the concentration dependency. The measured electrochemical redox current changed with the amount of the bound estradiol on the Au working electrode surface and the sensor can detect all the target material when the immobilized antibody amount is more than the estradiol amount in the water. The elecrochemical estradiol sensor without SAM treatment showed a low current of 7.79 nA, while the sensor treated with SAM resulted in 339 nA at 200 pg/ml, which is more than 40 fold higher output current. When combining the preconcentration technique and the SAM-treated electrode, the measured current became more than 100 fold higher than that of the sensor without neither SAM treatment nor preconcentration technique. The combination of these two techniques can would enable the proposed microfluidic electrochemical sensor to detect a very low concentration endocrine disruptor.

• Biotechnology and Bioprocess Engineering:BBE
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• v.11 no.2
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• pp.173-177
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• 2006

In this study, a novel strategy was developed for the highly selective immobilization of proteins, using the polyhydroxyalkanoate (PHA) depolymerase substrate binding domain (SBD) as an active binding domain. In order to determine the appropriacy of this method for immunodiagnostic assays, the single-chain antibody (ScFv) against the hepatitis B virus (HBV) preS2 surface protein and the severe acute respiratory syndrome coronavirus (SARS-CoV) envelope protein (SCVe) were fused to the SBD, then directly immobilized on PH A-coated slides via microspotting. The fluorescence-labeled HBV antigen and the antibody against SCVe were then utilized to examine specific interactions on the PHA-coated surfaces. Fluorescence signals were detected only at the spotted positions, thereby indicating a high degree of affinity and selectivity for their corresponding antigens/antibodies. Furthermore, we detected small amounts of ScFv-SBD (2.7 ng/mL) and SCVe-SBD fusion proteins (0.6ng/mL). Therefore, this microarray platform technology, using PHA and SBD, appears generally appropriate for immunodiagnosis, with no special requirements with regard to synthetic or chemical modification of the biomolecules or the solid surface.

• Archives of Pharmacal Research
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• v.21 no.3
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• pp.231-235
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• 1998

A sensitive enzyme immunoassay for serum TSH has been developed utilizing the tight binding between biotin and avidin, and three layered protein polystyrene beads as solid phase. To increase binding capacity of TSH and sensitivity of the assay, the polystyrene beads were coated sequentially with mouse immunoglobulin as first layer, rabbit antimouse immunoglobulin as second layer and monoclonal anti-TSH as third layer. A serum sample was incubated simultaneously with a monoclonal anti-TSH immobilized polystyrene beads and a second monoclonal anti-TSH covalently attached to biotin. After washing, the antibody bound serum TSH-anti-TSH-biotin complex is reacted with horseradish peroxidase (HRP)-labeled avidin. Following second wash, the bound HRP activity was measured calorimetrically. Reproducible results were obtained within 4 hours for serum TSH in the range between $0{\mu}\textrm{IU}$ml and ${50}{\mu}\textrm{IU}$ml with detection limit of $0.1{\mu}\textrm{IU}$ per test.

• Food Science and Biotechnology
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• v.14 no.6
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• pp.743-746
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• 2005

A dipstick-type immunochemical biosensor for the detection of the organophosphorus insecticide fenthion was developed using a screen-printed electrode system as an amperometric transducer with polyclonal antibodies against fenthion as a bioreceptor. The assay of the biosensor involved competition between the pesticide in the sample and pesticide-glucose oxidase conjugate for binding to the antibody immobilized on the membrane. This was followed by measurement of the activity of the bound enzyme by the supply of the enzyme substrate (glucose) and amperometric determination of the enzyme reaction product ($H_2O_2$). The activity of the bound enzyme was inversely proportional to the concentration of pesticide. The optimized sensor system showed a linear response against the logarithm of the pesticide concentration ranging from $10^{-2}$ to $10^3\;{\mu}g/L$.

• KSBB Journal
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• v.17 no.1
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• pp.1-13
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• 2002

Label-free optical methods for the monitoring of interactions between biological molecules have become increasingly popular within the last decade. A rising number of publications have demonstrated the benefits of direct biomolecular interaction analysis(BIA) for biology and biochemistry, such as antigen-antibody Interactions, receptor-ligand interactions, protein-DNA, DNA- intercalator, and DNA-DNA interactions. This article gives an overview of the historical development, principle and application of label-free optical biosensor to examine the functional characteristics of biospecific interaction, such as kinetics, affinity, and binding position of biomolecular between an immobilized species at the transducer surface and its dissolved binding partner.

• BMB Reports
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• v.31 no.5
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• pp.519-523
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• 1998

We have previously reported that anti-glucose-6-phosphate dehydrogenase (G6PD) serum raised against native G6PD (nG6PD) enzyme recognized nG6PD antigen poorly in competitive enzyme-linked immunosorbent assay (ELISA) (Kim, 1997). In the present study, we investigated whether anti-G6PD serum raised against nG6PD can react with denatured G6PD effectively in competitive ELISA. We used partially active G6PD (paG6PD) by repeated freeze-thawing or SDS-denatured G6PD (SDS-G6PD) as both immobilized and soluble antigens, and anti-G6PD serum raised against nG6PD for competitive ELISA. The polystyrene cuvettes coated with either paG6PD or SDS-G6PD were challenged with a mixture of a limiting amount of anti-G6PD serum and various doses of paG6PD or SDS-G6PD as competitors, followed by incubation with alkaline phosphatase-anti-IgG conjugate. The competitive ELISA with paG6PD or SDS-G6PD antigen exhibited the sigmoidal dose-response curve characteristic of competition immunoassays. Furthermore, Triton-denatured G6PD (Triton-G6PD) was used in competitive ELISA. The paG6PD, SDS-G6PD, or Triton-G6PD used as competitors increased the inhibition of antibody binding to immobilized either of nG6PD or denatured G6PD compared with nG6PD competitor. The inhibition by denatured G6PD competitors was more pronounced at high competitor concentrations than at low counterparts. We conclude that anti-G6PD serum raised against nG6PD can effectively react with denatured G6PD in competitive ELISA and that our anti-G6PD serum recognizes denatured enzymes better than active enzymes.

• Journal of Sensor Science and Technology
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• v.6 no.5
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• pp.356-361
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• 1997

Fiber-optic evanescent wave sensor was designed and fabricated to detect mouse immunoglobulin G(IgG) with decladed optical fiber on which anti-mouse IgG was immobilized. A sensitivity obtained by any direct or competitive method was lower than $1\;{\mu}g/m{\ell}$. Anti-mouse IgG was immobilized on 93.9% of core surface of optical fiber by simple adsorption method. The effect of postcoating using bovine serum albumin to remove non-specific binding was not observed. As the ratio of fluorescein to mouse IgG increased, the fluorescence signal increased, but that increase showed no linear relationship. Our fiber-optic sensor system could be used as immunosensor by measuring evanescent fluorescence in antigen-antibody reaction with good sensitivity below $1{\mu}g/m{\ell}$ level.