• Title/Summary/Keyword: 단백질칩

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Functional Expression of Anti-BNP scFv in E. coli Cytoplasm for the Detection of B-type Natriuretic Peptide (B-type natriuretic peptide 분석을 위한 항 BNP scFv 항체의 대장균 세포질 내에서의 기능적 발현)

  • Maeng, Bo-Hee;Nam, Dong-Hyun;Kim, Yong-Hwan
    • KSBB Journal
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    • v.24 no.6
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    • pp.591-597
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    • 2009
  • B-type natriuretic peptide is a neurohormone secreted in the cardiac ventricles. BNP levels are elevated in patients with ventricular dysfunction. Therefore, the concentration of BNP is important factor to reflect diagnosis and prognosis for cardiovascular disease. In this respect, anti-BNP scFv is an urgent requirement for early diagnosis in the field of biosensor. Herein, the genetic codes of anti-BNP scFv were chemically synthesized and cloned into both pET22b (+) and pColdⅣ vector, respectively. The recombinant scFv was successfully expressed as a functional form in cytoplasm of E. coli and detected through Western blot and ELISA. The highest level of functional expression of anti-BNP scFv was achieved using pET22b (+) vector at $15^{\circ}C$ by addition of 0.1 mM IPTG. Additionally, being exposed to both BNP and ANP, anti-BNP scFv specifically captured only BNP. Therefore, anti-BNP scFv expressed in this study will be applied to measure the concentration of BNP as a diagnostic recognition molecule.

다채널 표면 플라즈몬 공명 영상장치를 이용한 자기조립 단분자막의 표면 분석

  • Pyo, Hyeon-Bong;Sin, Yong-Beom;Yun, Hyeon-Cheol
    • 한국생물공학회:학술대회논문집
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    • 2003.04a
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    • pp.74-78
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    • 2003
  • Multi-channel images of 11-MUA and 11-MUOH self-assembled monolayers were obtained by using two-dimensional surface plasmon resonance (SPR) absorption. Patterning process was simplified by exploiting direct photo-oxidation of thiol bonding (photolysis) instead of conventional photolithography. Sharper images were resolved by using a white light source in combination with a narrow bandpass filter in the visible region, minimizing the diffraction patterns on the images. The line profile calibration of the image contrast caused by different resonance conditions at each points on the sensor surface (at a fixed incident angle) enables us to discriminate the monolayer thickness in sub-nanometer scale. Furthermore, there is no signal degradation such as photo bleaching or quenching which are common in the detection methods based on the fluorescence.

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Classification of Gene Data Using Membership Function and Neural Network (소속 함수와 유전자 정보의 신경망을 이용한 유전자 타입의 분류)

  • Yeom, Hae-Young;Kim, Jae-Hyup;Moon, Young-Shik
    • Journal of the Institute of Electronics Engineers of Korea CI
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    • v.42 no.4 s.304
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    • pp.33-42
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    • 2005
  • This paper proposes a classification method for gene expression data, using membership function and neural network. The gene expression is a process to produce mRNA and protains which generate a living body, and the gene expression data is important to find out the functions and correlations of genes. Such gene expression data can be obtained from DNA 칩 massively and quickly. However, thousands of gene expression data may not be useful until it is well organized. Therefore a classification method is necessary to find the characteristics of gene data acquired from the gene expression. In the proposed method, a set of gene data is extracted according to the fisher's criterion, because we assume that selected gene data is the well-classified data sample. However, the selected gene data does not guarantee well-classified data sample and we calculate feature values using membership function to reduce the influence of outliers in gene data. Feature vectors estimated from the selected feature values are used to train back propagation neural network. The experimental results show that the clustering performance of the proposed method has been improved compared to other existing methods in various gene expression data.

Interaction of Phenolic Compound-Specific Activator with Its Promoter using SPR-Based DNA Chip (SPR 근거 DNA 칩을 이용한 페놀 화합물 특이 CapR 조절 단백질과 촉진유전자와의 상호작용 연구)

  • 박선미;박후휘;임운기;신혜자
    • Journal of Life Science
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    • v.13 no.1
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    • pp.99-104
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    • 2003
  • Aromatic compounds are of major concern among environmental pollutants due to their toxicity and persistence. To monitor aromatic compounds in a real time with a better sensitivity, a new method of SPR (surface plasmon resonance) based on DNA chip (Biacore 3000) was developed here. It is thought that CapR regulatory protein as a complex with phenol, could bind to their corresponding promoter, Po. Biotinylated DNA oligomers for the promoter was synthesized by PCR and coupled onto streptoavidin-linked CM5-chip. CapR regulatory proteins were purified after cloning their genes in pET21a (+) vector and expressing proteins. The interaction was assessed by the system where the regulatory proteins flowed with or without phenol through the cells of DNA chip. CapR regulatory protein even in the presence of phenol had no response to its promoter, Po, suggesting that other factor(s) might be required for the activation of Po promoter. The present work reveals a promising possibility of the SPR-based DNA chip in monitoring specific environmental pollutants in a real time.

Injection Mold Technology of Protein Chip for Point-of-Care (현장진단용 단백질 칩 사출금형기술)

  • Lee, Sung-Hee;Ko, Young-Bae;Lee, Jong-Won;Jung, Hae-Chul;Park, Jae-Hyun;Lee, Ok-Sung
    • Design & Manufacturing
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    • v.6 no.2
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    • pp.74-78
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    • 2012
  • A multi-cavity injection mold system of protein chip for point-of-care with cavity temperature and pressure sensors was proposed in this work. In advance of manufacturing for the multi-cavity injection mold system, a single cavity injection mold system to mold protein chip was considered. Injection molding analysis for the presented system was performed to optimize the process of the molding and suggest guides to design. On the basis of the results for the single cavity system, a multi-cavity injection mold system for protein chip was analyzed, designed and manufactured with cavity temperature and pressure sensors. Results of balanced filling for protein chip models were obtained from the presented mold system.

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Investigation of the Binding Force between Protein A and Immunoglobulin G Using Dielectrophoretic(DEP) Tweezers Inside a Microfluidic Chip (미세유체 칩 내에서 유전영동 집게(Dielectrophoretic Tweezers) 를 이용한 단백질A와 면역 글로불린 G의 결합에 관한 연구)

  • Kwak, Tae Joon;Lee, Jae Woo;Yoon, Dae Sung;Lee, Sang Woo
    • Journal of Biomedical Engineering Research
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    • v.34 no.3
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    • pp.123-128
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    • 2013
  • The 'Dielectrophoretic Tweezers(DEP Tweezers)' can be used as a facile, economical toolkit for quantitative measurement of chemical and biological binding forces related to many biological interactions within a microfluidic device. Our experimental setup can probe the interaction between a single receptor molecule and its specific ligand. Immunoglobulin G(IgG) functionalized on polystyrene microspheres has been used to detect individual surface linked Staphylococcus protein A(SpA) molecules and to characterize the strength of the noncovalent IgG-SpA bond. It was measured and compared with the existing measurements. Measured single binding force of between Goat, Rabbit IgG and SpA were $17{\pm}7pN$, $74{\pm}16pN$. This work can be used to investigate several different ligand-receptor interactions and antigen-antibody interactions.

Gene Expression Profile in Epididymal Adipose Tissue from High-fat Diet Fed Mice (고지방식이를 섭취한 mouse에서의 유전자군의 발현양상 변화)

  • Cha, Min-Ho;Kim, Kyung-Seon;Sim, Woong-Seop;Yoon, Yoo-Sik
    • Korean Journal of Oriental Medicine
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    • v.8 no.2 s.9
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    • pp.75-84
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    • 2002
  • Obesity can be defined as a metabolic disease due to a increased state of fat tissue caused by an imbalance of calorie intake and use. To define genes that affected by different nutrient, we study gene expression from mice which were fed different nutrient. Epididymal and retro-peritineal adipose tissue were increase in high fat diet feeding mice compared with control, but liver and spleen were not. In serum, total cholesterol were differently increase in high fat diet feeding mice but total triglyceride and free fatty acid were not. That was maybe result of energy balance regulation in vivo system. aP2, PPART2 and FAS genes that were increased during adipogenesis were inclosed in high fat diet fed mice compared with control. In microarray assay, 1.4% of total genes were affected in epididymal adipose tissue by different nutrient. 1.1% of total genes were decreased down 0.5 fold and 0.3% were increased over 2 fold. These results indicated that many genes are affected in adipose tissue by nutrient.

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Fabrication of Paper-based Biosensor Chip Using Polydimethylsiloxane Blade Coating Method (PDMS 블레이드 코팅법을 이용한 종이-기반 바이오센서칩 제작)

  • Jeong, Heon-Ho;Park, Chami
    • Korean Chemical Engineering Research
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    • v.59 no.1
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    • pp.100-105
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    • 2021
  • This paper proposes the polydimethylsiloxane (PDMS) blade coating method for fabrication of paper-based analytical device (PAD) that is able to monitor the disease diagnosis and progress without special analytical equipment. The mold that has PAD design is easily modified by using laser cutting technique. And the fabricated mold is used for hydrophobic barrier formation by blade coating. We have optimized the stable formation of PDMS hydrophobic barrier as blade coating condition, which is established by analyzing the structure of the PDMS hydrophobic barrier and change of hydrophilic channel size as thickness of the ink and contact time with the chromatography paper. Based on optimal condition, we demonstrate that PAD as biosensor can apply to detect protein, glucose, and metal ion without special analysis equipment.

Development of Lateral Flow Immunofluorescence Assay Applicable to Lung Cancer (폐암 진단에 적용 가능한 측면 유동 면역 형광 분석법 개발)

  • Supianto, Mulya;Lim, Jungmin;Lee, Hye Jin
    • 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.

Optimization of SELDI-TOF MS for Peptide Profiling of Sorghum Seed (수수종자의 펩타이드 분석을 위한 SELDI-TOF MS 최적화 연구)

  • Park, Sei Joon;Park, June Young;Lee, Yong Ho;Hwang, Su Min;Kim, A Ram;Ko, Jee-Yeon;Kim, Tae Wan
    • KOREAN JOURNAL OF CROP SCIENCE
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    • v.58 no.1
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    • pp.50-56
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    • 2013
  • For accurate analysis of low molecular peptides using SELDI-TOF MS (surface enhanced laser desorption/ionization time of flight mass spectrometry), the optimized analytical conditions should be established for a specific biological sample. This study was conducted to optimize SELDI-TOF MS analytical conditions for profiling low molecular peptide below 10 kDa presented in sorghum seeds. Analytical conditions were as follows; (1) protein chips: CM10 (weak cation exchanger) and Q10 (strong anion exchanger), (2) dilution factors of binding buffer: 1/2, 1/5, 1/10, 1/20, 1/50, 1/100, and 1/200, (3) the stringency of Q10 binding buffer: 10 mM and 100 mM, and (4) protein extraction buffers: sodium borate, sodium borate + acetone, phenol, and TCA buffers. Optimum dilution factors were selected as 1/20 and 1/50 in both protein chips, CM10 and Q10. Low stringency of Q10 binding buffer (10mM) detected more peptide peaks than high stringency (100 mM). Selected protein extraction buffers of sorghum seed for SELDI-TOF MS analysis was the sodium borate buffer in the range of 2~10 kDa, while the phenol buffer was more suitable in the range of 10~20 kDa.