• 센서학회지
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• 제14권5호
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• pp.308-312
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• 2005

We present a microfluidic chip designed for the detection of antibody by using quantum dots fluorescence and a microbead-based assay. A custom designed PDMS microfluidic chip with multi-layer channel is utilized for capturing microbeads; antibody injection into each micro-well; QD injection; and fluorescence detection. The experiment using the fabricated microfluidic chip has been performed on solutions with various concentrations of antibody and has shown correlated fluorescent intensities.

• 한국미생물·생명공학회지
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• 제17권3호
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• pp.269-272
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• 1989

In order to obtain monoclonal antibody from ascites fluid at sufficiently high purity using a single hydroxylapatite chromatography (HA) a further optimization on its operating variables was carried out. By adjusting the pH of the eluent, the sodium phosphate buffer, to 6.0 from 6.8 and adding CaCl$_2$to 1 mM at the column inlet, the elution molarities (M$_{elu}$) for the desired monoclonal antibody and contaminating proteins can be distinguished from each other with enough resolution. Previously these two groups of proteins co-eluted at the same time at pH 6.8 and without CaCl$_2$. This sin81e step hydroxylapatite chromatography yields the desired antibody pure enough for diagnostic use.

• 산업식품공학
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• 제14권1호
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• pp.21-26
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• 2010

C-Reactive protein (CRP), which is an 118 kDa pentameric protein, was secreted by the liver is an important biomarker for coronary disease, hypertension and inflammation. In this study, a method for CRP detection exploiting quantum dot (Qdot)-antibody conjugate was developed according to an indirect-competitive immunosensing protocol. For this purpose, a streptavidin-bound $Qdot_{605}$ was linked with a separately prepared biotinylated monoclonal antirat CRP antibody to produce a Qdot-antibody conjugate. The immunosensing was performed at 0.1 and 20 nM of the coating antigen and conjugate, respectively. The current method was found very sensitive in CRP detection, judging from the concentration-dependent fluorescence emission.

• 산업식품공학
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• 제13권4호
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• pp.314-319
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• 2009

Metallothioneins (MTs) are low molecular weight, cysteine-rich, and heavy metal binding proteins, which could be induced with heavy metals such as Cd, Hg, Zn and Cu in liver, kidney, and in cultured cells. By using ion exchange chromatography on DE-52, MT was purified from the serum of carp induced with cadmium in order to produce antibody against MT. Polyclonal antibody produced against purified carp MT reacted well with MT in the serum of carp induced with cadmium, whereas control serum did not. This may indicate that the polyclonal antibody against the carp MT could be used for the preparation of biosensors to detect MT in fishes like carp.

• Asian Pacific Journal of Cancer Prevention
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• 제16권12호
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• pp.5115-5118
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• 2015

Background: The $p16^{INK4a}$ is a protein that expressed in Liquid-based cervical cytology specimens and has been proved link to cervical cancer. The $p16^{INK4a}$ could be detection by piezoelectric immunosensor and the immobilization of the $p16^{INK4a}$ antibody influence the sensitivity of the piezoelectric immunosensor. Materials and Methods: $5{\mu}L$ mouse polyclonal antibody against $p16^{INK4a}$ was bound onto the surface of immonosensor through two methods. (directly immobilized method; protein A method). Absorb of the $p16^{INK4a}$ antibody on the surface of immonosensor caused a shift in the resonant frequency of the immunosensor and The frequency changes recorded showed a better reproducibility. The activity of the immobilization antibody with the directly method and protein A method was tested with $p16^{INK4a}$ antigen. Results: The resonant frequency for different antibody immobilization methods were different, and the sensitivity for $p16^{INK4a}$ detection also different. Conclusions: The protein A method was found to be much more better than the directly method for the immobilization of the p16INK4A antibody on the gold electrode of the quartz crystal for cervical lesion detection. The Protein A method created more reproducible and stable immobilization antibody layers with p16INK4A antigen.

• BMB Reports
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• 제33권1호
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• pp.92-95
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• 2000

The HIV-1 p24(202-221) sequence ETINNEEEWDRVHPV HAGP contains a B-cell epitope with the earliest immune response and the highest antibody titer against anti-mouse sera obtained by immunization with p24 antigens. A novel mouse monoclonal antibody (mAb) was generated against the immunodominant B-cell epitope of the HIV-1 p24 capsid protein, p24(202-221). BALB/c mice were immunized with the four branched multiple antigenic peptide (MAP) containing the HIV-1p24(202-221) sequence, and antibody-secreting hybridoma were produced by fusion of mouse splenocytes with P3X63Ag8.653, mouse myeloma cells. One clone which produced the antigen-specific mAb named KI-24 (Isotype IgG1, light chain: ${\kappa}$) was identified. mAb KI-24 was highly specific for both the p24(202-221) and p24 proteins when analyzed by ELISA and Western blotting. Since p24(202-221) also contains a cytotoxic T-lymphocyte epitope, this specfic peptide epitope and the monoclonal antibody with specific reactivity against the p24 protein and p24(202-221) can be used in peptide vaccine development and p24 antigen detection from HIV patients.

• 센서학회지
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• 제19권4호
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• pp.306-312
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• 2010

We have detected deoxynivalenol(DON) using a metal-oxide-semiconductor field-effect-transistor(MOSFET)-based biosensor. The MOSFET-based biosensor is fabricated by a standard complementary metal-oxide-semiconductor(CMOS) process, and the biosensor's electrical characteristics were investigated. The output of the sensor was stabilized by employing a reference electrode that applies a fixed bias to the gate. Au which has a chemical affinity for thiol was used as the gate metal to immobilize a self-assembled monolayer(SAM) made of 16-mercaptohexadecanoic acid(MHDA). The SAM was used to immobilize anti-deoxynivalenol antibody. The carboxyl group of the SAM was bound to the anti- deoxynivalenol antibody. Anti-deoxynivalenol antibody and deoxynivalenol were bound by an antigen-antibody reaction. In this study, it is confirmed that the MOSFET-based biosensor can detect deoxynivalenol at concentrations as low as 0.1 ${\mu}g$/ml. The measurements were performed in phosphate buffered saline(PBS; pH 7.4) solution. To verify the interaction among the SAM, antibody, and antigen, surface plasmon resonance(SPR) measurements were performed.

• Biotechnology and Bioprocess Engineering:BBE
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• 제4권1호
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• pp.72-77
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• 1999

An antibody containing a genetically engineered lipid group at the N-termunus and a hexahistidinyl tag at the C-terminus (Lpp-scF-His6) was immobilized in an oriented manner on the surface of liposome. Liposomes, consisting of antibody and phosphatidyl-choline, have been prepared and imaged by AFM. For AFM visualization, the resulting liposomes were bound on the surface of mica by two different mechanisms. The histidine tags present in the antibody molecules of the immonuliposome were anchored to the NiCl2 treated mica surface. Alternatively, the immunoliposomes were immunochemically bound on antigen-coated mica surface. Both approaches yielded liposomes which were clearly imaged without damage by AFM in ambient condition.

• 한국생물공학회:학술대회논문집
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• 한국생물공학회 2002년도 생물공학의 동향 (X)
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• pp.293-296
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• 2002

Hyperosmotic pressure increased specific antibody productivity ($q_{Ab}$) of recombinant CHO cells (SH2-0.32) while it depressed cell growth. Thus, the use of hyperosmolar medium did not increase the maximum antibody concentration substantially. To overcome this drawback, the feasibility of biphasic culture strategy was investigated. In the biphasic culture, cells were first cultivated in the standard medium with physiological osmolality(294 mOsm/kg) for cell growth. When cells reached the late exponential phase of growth, the spent standard medium was replaced with the fresh hyperosmolar medium (522 mOsm/kg) for antibody production. The ($q_{Ab}$) in growth phase with the standard medium was 2.1 ${\mu}g/10^6cell/day$ while the ($q_{Ab}$) in antibody production phase with the hyperosmolar medium (522 mOsm/kg) was 11.1 ${\mu}g/10^6cell/day$. Northern blot analysis showed a positive relationship between the relative contenet of Ig mRNA and ($q_{Ab}$), indicating that transcriptional regulation was involved in the response of rCHO cells to hyperosmotic pressure. Due to the enhanced ($q_{Ab}$) and increased cell concentration in biphasic culture, the maximum antibody concentration obtained in biphasic culture with 522 mOsm/kg medium exchange was 161% higher than that obtained in batch culture with the standard medium. Taken together, simple biphasic culture strategy based on hyperosmotic culture for improved foreign protein production from rCHO cells is effective in improving antibody production of rCHO cells.

• IMMUNE NETWORK
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• 제1권3호
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• pp.187-195
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• 2001

The expression of recombinant proteins fused to 26 kDa glutathione S-transferase (GST) extracted from Schistosoma japonicum represents an attractive system for purifiying proteins of interest in a single step using GST-affinity chromatography. In addition, the GST-tag is used conveniently for detecting fused proteins since its high solubility as well as its relatively small size rarely interferes with the biological activity of the fused protein. In this regard, the GST system is frequently applied for tracing fusion proteins in both prokaryotic and eukaryotic cells to elucidate the physiological interactions and functional compartments of proteins. To provide a further tool in analyzing GST-fusion proteins, a new monoclonal antibody, with a high specificity to the S. japonicum GST was produced. Methods: BALB/c mice were immunized both with recombinant S. japonicum GST proteins, and by the fusion of splenocytes from these mice with myeloma cells. From this, a new anti -GST monoclonal antibody, termed SARAH, was generated. The specificity and reactivity of this antibody was confirmed by ELISA and by Western blot analysis. Results: SARAH showed a high reactivity to recombinant GST and GST fusion protein but not with native mammalian GST proteins as derived from other species including humans, cows, rabbits and rats. The applicability of SARAH was further demonstrated by confocal laser scanning microscopy, where GST proteins that were expressed transiently in mouse fibroblast cells, were specifically detected without interference of endogenous GST. Conclusion: SARAH is new monoclonal antibody with a high specificity to recombinant GST proteins but not to endogenous GST in mammalian cells.