• Journal of Photoscience
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• 제10권3호
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• pp.237-240
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• 2003

The tetradecameric peptide (K47-K60) near the NH$_2$-terminal region of epithelial-cell adhesion molecule (Ep-CAM) was chosen as antigenic site and a polyclonal antibody was generated, which could recognize Ep-CAM from the mouse colon tissue or the colon cancer cell, CT-26, in Western blot analysis. Then, the fluorescein isothiocyanate (FITC), a fluorescence dye, was conjugated with the affinity purified Ep-CAM antibody using thiocyanate and the amino groups of FITC and antibody, respectively. The molar ratio of FITC to antibody was estimated approximately 1.86 to 1.00 by measuring the optical densities at 492 nm and 280 nm. Ep-CAM antibody-FITC conjugate was then used for immunohistochemistry of the CT-26 cells. Judging from the shapes formed by fluorescence, the Ep-CAM antibody could delivered FITC to the surface of cells in which Ep-CAM was expressed. This result implies that Ep-CAM antibody could be also used for the tissue-specific delivery of the photosensitizer to the target protein via antigen-antibody interaction.

• 한국식품영양학회지
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• 제8권2호
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• pp.70-78
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• 1995

We had examined the levels of specific IgG and IgA to dietary antigens in human breast milk and the relationships between the maternal food intake and the specific antibody level. The highest antibody titers were found in colostrum and decreased as lactation progressed. The specific antibody level was not affected by maternal calorie or protein intake, but affected by the intake frequency of a kind of food. Egg and meat intake significantly related to anti-OVA IgG and anti-BSA IgA antibodies, respectively. Meat intake frequency was generally affected by the other specific antibody levels.

• Journal of Photoscience
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• 제12권3호
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• pp.123-127
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• 2005

The polyclonal antibody was generated against the peptide fragment of 62 amino acid residues (D 181-T242) near the COOH-terminal region of the extracellular domain of epithelial-cell adhesion molecule (Ep-CAM) and shown to be able to recognize Ep-CAM in competitive ELISA. Then, sulforhodamine 101 acid chloride (so called Texas red), a fluorescence dye, was conjugated to the affinity-purified anti-Ep-CAM antibody utilizing the reaction between the aliphatic amines of antibody and the sulfonyl chloride of Texas red. The molar ratio of Texas red to antibody was estimated to be approximately 1.86 by measuring optical densities at 280 nm and 596 nm, implying that the two molecules of Texas red at most were conjugated to antibody. The anti-Ep-CAM antibody-Texas red conjugate was then used for immunohistochemistry of CT-26 murine colon carcinoma cells. Based upon the fluorescence microscope images, anti-Ep-CAM antibody is able to deliver Texas red specifically to the surface of CT-26 cells on which Ep-CAM was actively expressed. This result indicates that anti-Ep-CAM antibody could be useful for the tissue-specific delivery of photosensitizers via antigen-antibody interaction.

• 생약학회지
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• 제19권2호
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• pp.127-132
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• 1988

In the course of the immunological studies on the natural products, the antibody specific to saikosaponin a was producted. Saikosaponin a was treated with succinic anhydride to give 6'-O-hemisuccinyl saikosaponin a, which was successively converted to saikosaponin a-BSA conjugate (4. 5 mole saikosaponin a/mole of BSA) by carbodiimide method. The antibody obtained from rabbits immunized with saikosaponin a-BSA conjugate as usual manner reacted with both the conjugate and BSA, while after the absorption with BSA, the antibody reacted with the conjugate alone.

• Parasites, Hosts and Diseases
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• 제37권4호
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• pp.243-248
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• 1999

The present study analyzed serum IgG subclass antibody reaction to major antigenic bands of Clonorchis sinensis to investigate improvement of its serodiagnosis. Of the four subclass antibodies, IgG1 and IgG2 antibodies were produced but not specific, IgG3 antibody was least produced, and IgG4 antibody was prominent and specific. The serum IgG antibody reaction to any of 43-50, 34-37, 26-28, and 8 kDa bands was found in 65.5％ of 168 egg positive cases while IgG4 antibody reaction was found in 22.0％ of them. The positive rates of IgG and IgG4 antibodies were directly correlated with the intensity of infection. All of the sera from heavily infected cases over EPG 5,000 showed positive reaction for specific IgG and IgG4 antibodies. The specific serum IgG4 antibody disappeared within 6 months after treatment. The bands of 35 kDa and 67 kDa cross-reacted with IgG antibodies but not with IgG4 antibodies in sera of other trematode infections. The present findings suggest that serum IgG4 antibody reaction to 8 kDa band is specific but not sensitive. Any method to increase its sensitivity is required for improved serodiagnosis.

• Molecules and Cells
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• 제27권3호
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• pp.313-319
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• 2009

The dual-vector system-II (DVS-II), which allows efficient display of Fab antibodies on phage, has been reported previously, but its practical applicability in a phage-displayed antibody library has not been verified. To resolve this issue, we created two small combinatorial human Fab antibody libraries using the DVS-II, and isolation of target-specific antibodies was attempted. Biopanning of one antibody library, termed DVFAB-1L library, which has a $1.3{\times}10^7$ combinatorial antibody complexity, against fluorescein-BSA resulted in successful isolation of human Fab clones specific for the antigen despite the presence of only a single light chain in the library. By using the unique feature of the DVS-II, an antibody library of a larger size, named DVFAB-131L, which has a $1.5{\times}10^9$ combinatorial antibody complexity, was also generated in a rapid manner by combining $1.3{\times}10^7$ heavy chains and 131 light chains and more diverse anti-fluorescein-BSA Fab antibody clones were successfully obtained. Our results demonstrate that the DVS-II can be applied readily in creating phage-displayed antibody libraries with much less effort, and target-specific antibody clones can be isolated reliably via light chain promiscuity of antibody molecules.

• IMMUNE NETWORK
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• 제3권3호
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• pp.219-226
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• 2003

Background: Phage display is the most widely used technique among display methods to produce monoclonal antibody fragment with a specific binding activity. Having a large library for efficient antibody display/selection is quite laborious process to have more than $10^9$ members of transformants. To overcome these limitations, several in vitro selection approaches have been reported. Ribosome display that links phenotypes, proteins, directly to genotype, mRNA, is one of the in vitro display methods. Ribosome display can reach the size of scFv library up to $10^{14}$ molecules and it can be further diversified during PCR steps. To select the high affinity scFv from one pot library, we established ribosome display technique by modifying the previously reported eukaryotic translation system. Methods: To establish the antibody selection system by ribosome display, we used 3D8, anti-DNA antibody. A 3D8 scFv was synthesized in vitro by an in vitro transcription-translation system. The translated 3D8 scFv and the encoding 3D8 mRNA are connected to the ribosome. These scFv-ribosome-mRNA complexes were selected by binding to their specific antigens. The eluted mRNAs from the complexes are reverse transcribed and re-amplified by PCR. To apply this system, antibody library from immunized mouse with terminal protein (TP)-peptide of hepatitis B virus DNA polymerase TP domain was also used. This TP-peptide encompasses the 57~80 amino acid residues of TP. These mRNA/ribosome/scFv complexes by our system were panned three times against TP-peptide. The enrichment of antibody from library was determined by radioimmunoassay. Results: We specifically selected 3D8, anti-DNA antibody, against ssDNA as a model system. The selected 3D8 RNAs sequences from translation complexes were recovered by RT-PCR. By applying this model system, we enriched TP-peptide-specific scFv pools through three cycles of panning from immunized library. Conclusion: We show that our translating ribosome complexes are well maintained and we can enrich the TP-specific scFv pools. This system can be applied to select specific antibody from an antibody library.

• BMB Reports
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• 제38권6호
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• pp.646-649
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• 2005

Human neuronal growth inhibitory factor (GIF), a metalloprotein classified as metallothionein-3, is specifically expressed in mammal central nervous system (CNS). In these Studies the specific antibody to human GIF was prepared and used to search the epitope of human GIF by enzyme-linked immunosorbent assay (ELISA) and sequence comparison. The result of ELISA showed the epitope of human GIF may locate on a octapeptide (EAAEAEAE) in the $\alpha$-domain of human GIF, and the result of nerve cell culture indicated that the biological activity of GIF may be affected by the specific antibody.

• 대한수의학회지
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• 제29권1호
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• pp.27-35
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• 1989

To develop more specific and sensitive diagnostic methods for infectious bovine rhinotracheitis, 7-C-2 monoclonal antibody specific to polypeptides of infectious bovine rhinotracheitis virus (IBRV) was applied in indirect immunofluorescence antibody assay (IFA), indirect immunoperoxidase assay(IPA) and radial immunodiffusion enzyme assay (RIDEA). It was found that IBRV infected in MDBK cells could be detected as early as 8 hours post infection by IFA, and that IFA was more rapid and specific to identify IBRV antigen than IPA. The diagnostic efficacy of RIDEA and SN test was studied with 88 bovine sera. It was evident that RIDEA could eliminate the false positive reaction encountered in serum neutralization(SN) test, being more rapid and sensitive than the latter. Highly significant correlation coefficiency (r=0.76, p<0.01) was evaluated between the titers of sera and the diameters of RIDEA. Tracheal membranes and sera collected from 96 slaughtered cattle with lesions in respiratory organs were examined to detect IBRV antigen and antibody by IFA, RIDEA and SN test. It was presented that positive rates were 32.3% in IFA, 20.8% in RIDEA and 21.9% in SN test, and that coincidence rate between RIDEA and SN test were 100% in positive sera and 98.7% in negative sera. In conclusion, it was assumed that application of monoclonal antibody could improve the diagnostic efficacy of IBR by enhancing sensitivity and specificity of IPA, IFA and RIDEA.

• 대한핵의학회지
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• 제27권2호
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• pp.261-269
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• 1993

핵의학 분야에 사용되는 항체의 적절한 동위원소 표지법을 선택하는데는 표지항체의 면역반응성을 평가하는 것이 매우 중요한 일이다. 저자 등은 국내에서 개발된 CEA-79 단세포군 항체를 표지방법 및 비방사능을 $0.1{\mu}Ci/{\mu}g$으로부터 $100{\mu}Ci/{\mu}g$까지 달리하여 $^{125}I$ 표지를 시행한 후 면역반응성을 평가하고, 이 항체에 적절한 표지방법을 찾고자 하였다. 면역반응성의 평가에 있어서는 표지, 비표지 항체의 경쟁반응을 이용하여 표지항체의 비표지항체에 대한 상대적 면역반응성을 평가하는 새로운 방법을 시도해 보았다. 그 결과 CEA-79 항체의 강우, chlormine T법이 iodogen법보다 우수하였는데, 즉, chloramine T법으로 표지시에는 비방사능을 $100{\mu}Ci/{\mu}g$까지 높여도 면역반응성이 잘 유지되고 시간경과에 따른 면역반응성의 변화도 적었다. 본 연구에서 시도한 새로운 경쟁반응방법은 동위원소표지에 의한 항체의 면역반응성의 손상을 평가하는데 효과적이었으며, 기존의 세포결합 검사법보다 간편하였다. 또 항체의 면역반응성은 비방사능과 함께 방사면역계수법의 민감도에 영향을 미침을 확인할 수 있었다. 따라서 표지항체의 면역반응성과 비방사능은 그 용도에 따라 적정화되어야 할 것이며, 본 연구에서 시도한 경쟁반응방법은 적정한 동위원소 표지 방법을 선택하는데 유용할 것으로 사료된다.