• BMB Reports
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• v.31 no.1
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• pp.101-105
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• 1998

Immunoassay of botulinum toxin (BTX) B type was investigated using two typed of biosensors: light addressable potentiometric sensor (LAPS) and surface plasmon resonance (SPR) sensor. Urease-tagged and immuno-filtration capture method have been used for LAPS. Tag-free and direct binding real-time detection method have been used for SPR sensor. The detection limit of sandwich assay format with LAPS was 10 ng/ml, which was the lowest among methods tested. SPR has the advantage of being more convenient because tag-free direct binding assay can be used and reaction time was reduced, regardless of low sensitivity. This result shows that sandwich assay format with LAPS can be used as an alternative method of BTX mouse bioassay which is known as the most sensitive method for the detection of BTX.

• Applied Chemistry for Engineering
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• v.35 no.4
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• pp.335-340
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• 2024

In this paper, we synthesized organic and inorganic hybrid materials to introduce antibody functionality to UIO-66 and incorporated them into a surface plasmon resonance (SPR) assay to enhance the sensitivity of detecting small molecules such as oxytocin. A biological marker peptide called oxytocin may help in the diagnosis of heart failure, Alzheimer's disease, and cancer. To detect oxytocin at concentrations as low as a few femtomole (fM), we developed a surface sandwich assay utilizing a pair of oxytocin-specific antibodies for enhancing selectivity and one of metal organic frameworks [e.g., UiO-66-(COOH)₂] possessing high porosity and surface-area as a signal amplifier. Initially, real-time SPR assays were used to confirm that each selected oxytocin-specific antibody binds strongly to oxytocin and to different binding sites on oxytocin. One of these antibodies (e.g., anti-OXT[OTI5G4]) was immobilized on the surface of a thin gold chip. Upon sequential injecting of oxytocin and the other antibody (e.g., anti-OXT[4G11]) conjugated to UiO-66-(COOH)₂ onto the surface to form the surface sandwich complex of anti-OXT[OTI5G4]/oxytocin/UiO-66-(COOH)₂-anti-OXT[4G11]), SPR changes, which varied with oxytocin concentration, were then measured in real time. The results demonstrated that sensitivity was amplified by over a million-fold compared to assays without UiO-66-(COOH)₂, enabling oxytocin detection down to approximately 10 fM.

• Journal of Sensor Science and Technology
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• v.16 no.6
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• pp.441-448
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• 2007

In this study, the fiber-optic fluoro-immunosensor was designed to detect foodborne pathogens. The fabricated system is composed of the multimode optical fiber on which antibodies are immobilized. Then, a sandwich immunoassay is applied to the fabricated the fiber-optic fluoro-immunosensor. In the "sandwich" binding format, a primary or "capture" antibody is immobilized on the core surface of the multimode optical fiber and a secondary or named as "tracer" antibody is added to the bulk solution. A tracer is labeled FITC (fluorescein isothiocyanate; ${\lambda}ex$=492 nm, ${\lambda}em$= 520 nm). Different concentrations of antigens are tested in different fibers. The detection limit of the fabricated system is 5.08×103 cfu/ml for Vibrio antigen and $0.1{\mu}g/ml$, $0.05{\mu}g/ml$ in non-labeled monolayer phosphate buffered saline (NMP), non-labeled monolayer carbonate bicarbonate buffer (NMC), respectively.

• KSBB Journal
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• v.11 no.4
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• pp.420-430
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• 1996

One-step immuno-chromatography assay system for heat-killed Salmonella typhimurium antigens was developed. Three major components used were a glass fiber membrane (placed at the bottom of the system) with an antibody (specific to the analyse, detection antibody)-gold conjugate deposited in a dry state on the surface, a nitrocellulose membrane (middle) with an antibody (also, specific to the analyse but recognized different epitome: capture antibody) and anti-detection antibody immobilized in spatially separated areas, and a cellulose membrane (top) as absorption pad. These membranes were partially superimposed such that a wicking of aqueous solution containing sample can continuously take place through membranes. Variables that affected the system performance were the concentration of capture antibody, the location on the membrane, inert protein used for blocking of the membrane and for carrying the sample, and the concentration of the gold conjugate. Under optimal conditions, within 15 minutes after absorption of a sample solution from the bottom of the system antigen-antibody complexes of sandwich type were formed on the membrane surface area with immobilized capture antibody and a color signal was generated in proportion to the analyse concentration. The minimum do tection limit of the analyse was $1{\times}106$ Salmonella cells/mL.

• 한국생물공학회:학술대회논문집
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• 2005.04a
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• pp.47-47
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• 2005

There are many different surface technologies currently applied for preparation of protein chips. However, it requires innovative surface chemistry for capture proteins to be immobilized on chip surface keeping their conformation and activity intact and their orientation right, while they bind tightly and densely in a given array spot. Proteogen has developed 'ProteoChip BP' coated with novel proprietary linker molecules $(ProLinker^{TM})$ for efficient and robust immobilizations of capture proteins by improving surface properties of molecular captures. It was demonstrated that $ProLinker^{TM}$ gave the best surface performance in preparation of protein microarray chip base plates among others currently available on the market. In particular, the $ProLinker^{TM}-based$ surface chemistry has demonstrated to provide excellent performance in preparation of 'Antibody Chip' for analysis of biomarkers as well as proteome expression profiles. The linker molecule has also shown to be well applicable for development of biosensors and micro-beads as well as protein microarray and nano-array. ProteoChip BP can be used either for preparation of high-density array by using a microarrayer or for preparation of 'Well-on-a-Chip' with low density array, which is better applicable for quantitative analysis of biomarkers or protein-protein interactions. The biomarker assay can be performed either by direct or sandwich methods of fluorescence immunoassay. Application of ProteoChip BP has been well demonstrated by the extensive studies of 1) tumor-marker assays, 2) new drug screening by using 'Integrin Chip' and 3) protein expression profile analysis. Some of experimental results will be presented.

• Proceedings of the Plant Resources Society of Korea Conference
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• 2020.12a
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• pp.3-13
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• 2020

Pseudomonas aeruginosa is a ubiquitous gram-negative bacterium causing serious infections. The P. aeruginosa T3SS is a syringe-like apparatus on the bacterial surface, with 4 effector toxins: ExoS, ExoT, ExoY, and ExoU. Here, we investigated the effect of ExoS and ExoT of the T3SS of P. aeruginosa K strain (PAK). The type three secretion system (T3SS) is a major virulence system of Pseudomonas aeruginosa (P. aeruginosa). The effector protein Exotoxin S (ExoS) produced by P. aeruginosa is secreted into the host cells via the T3SS. For the purpose of screening the inhibitors with regard to ExoS secretion, we developed the sandwich-type enzyme-linked immunosorbent assay (ELISA) system. PAK clinical strains induce proinflammatory cytokine production through the T3SS, and this involves NF-κB activation in pneumonia mouse models. We tried to confirm the role of the NF-κB transcription factor in ExoS- and ExoT-induced pneumonia mouse models. pro-inflammatory cytokines induction in response to ExoS and ExoT infection relied on NF-κB activation. Our findings highlight the roles of natural poduct in inhibiting proinflammatory cytokine expression during ExoS and ExoT exposure in PAK infections, paving the way for a novel therapeutic approach for the treatment of pulmonary infections.

• Applied Chemistry for Engineering
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• v.28 no.4
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• pp.397-403
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• 2017

The number of patients suffering from Alzheimer's disease is increasing year after year and almost approaching 15% of the total elderly population. Although it is critical to detect the early stage of Alzheimer's disease, which is a serious illness causing cognitive deficits, various existing diagnosis methods such as MRI, PET and CSF analysis could be the burdens for patients due to their high costs and long time to diagnosis. In order to tackle some of challenging issues for such existing diagnosis methods, extensive efforts have been made on developing fast and convenient biochip sensing methodologies for the diagnosis of Alzheimer's disease with a droplet of patient biofluids (e.g., blood). In this mini-review, we highlight some of the latest biochip sensing technologies that could qualitatively and quantitatively analyze blood biomarkers used for Alzheimer's disease diagnostics and discuss briefly related research trends and future aspects.

• Macromolecular Research
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• v.14 no.5
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• pp.565-572
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• 2006

Recombinant human bone morphogenic protein-2 (rhBMP-2), which is known as one of the major local stimuli for osteogenic differentiation, was immobilized on the surface of hyaluronic acid (HA)-modified poly$(\varepsilon-caprolactone)$ (PCL) (HA-PCL) scaffolds to improve the attachment, proliferation, and differentiation of human bone marrow stem cells (hBMSCs) for bone tissue engineering. The rhBMP-2 proteins were directly immobilized onto the HA-modified PCL scaffolds by the chemical grafting the amine groups of proteins to carboxylic acid groups of HA. The amount of covalently bounded rhBMP-2 was measured to 1.6 pg/mg (rhBMP/HA-PCL scaffold) by using a sandwich enzyme-linked immunosorbant assay. The rhBMP-2 immobilized HA-modified-PCL scaffold exhibited the good colonization, by the newly differentiated osteoblasts, with a statistically significant increase of the rhBMP-2 release and alkaline phosphatase activity as compared with the control groups both PCL and HA-PCL scaffolds. We also found enhanced mineralization and elevated osteocalcin detection for the rhBMP-2 immobilized HA-PCL scaffolds, in vitro.

• Journal of Sensor Science and Technology
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• v.23 no.4
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• pp.278-283
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• 2014

In this study, we propose an immunosensor using zinc oxide nanorods (NRs) inside PDMS channel for detecting the influenza A virus subtype H7N9. ZnO with high isoelectric point (IEP, ~9.5) makes it suitable for immobilizing proteins with low IEP. In this proposed H7N9 immunosensor structure ZnO NRs were grown on the PDMS channel inner surface to immobilize H7N9 capture antibody. A sandwich enzyme-linked immunosorbent assay (ELISA) method with was used 3,3',5,5' tetramethylbenzidine (TMB) for detecting H7N9 influenza virus. The immunosensor was evaluated by amperometry at various H7N9 influenza antigen concentrations (1 pg/ml - 1 ng/ml). The redox peak voltage and current were measured by amperometry with ZnO NWs and without ZnO NWs inside PDMS channel. The measurement results of the H7N9 immunosensor showed that oxidation peak current of TMB at 0.25 V logarithmically increased from 2.3 to 3.8 uA as the H7N9 influenza antigen concentration changed from 1 pg/ml to 1 ng/ml. And then we demonstrated that ZnO NRs inside PDMS channel can improve the sensitivity of immunosensor to compare non-ZnO NRs inside PDMS channel.

• Applied Chemistry for Engineering
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• v.33 no.2
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• pp.173-178
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• 2022

A lateral flow immunoassay (LFIA) method using carbon nanodot@silica as a signaling material was developed for analyzing the concentration of retinol-binding protein 4 (RBP4), one of the lung cancer biomarkers. Instead of antibodies mainly used as bioreceptors in nitrocellulose membranes in LFIA for protein detection, aptamers that are more economical, easy to store for a long time, and have strong affinities toward specific target proteins were used. A 5' terminal of biotin-modified aptamer specific to RBP4 was first reacted with neutravidin followed by spraying the mixture on the membrane in order to immobilize the aptamer in a porous membrane by the strong binding affinity between biotin and neutravidin. Carbon nanodot@silica nanoparticles with blue fluorescent signal covalently conjugated to the RBP4 antibody, and RBP4 were injected in a lateral flow manner on to the surface bound aptamer to form a sandwich complex. Surfactant concentrations, ionic strength, and additional blocking reagents were added to the running buffer solution to optimize the fluorescent signal off from the sandwich complex which was correlated to the concentration of RBP4. A 10 mM Tris (pH 7.4) running buffer containing 150 mM NaCl and 0.05% Tween-20 with 0.6 M ethanolamine as a blocking agent showed the optimum assay condition for carbon nanodot@silica-based LFIA. The results indicate that an aptamer, more economical and easier to store for a long time can be used as an alternative immobilizing probe for antibody in a LFIA device which can be used as a point-of-care diagnosis kit for lung cancer diseases.