• Food Science and Biotechnology
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• 제18권2호
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• pp.317-322
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• 2009

The proteins from 15 types of cultivar of hot peppers cultivated in Korea were extracted and its allergenicity was investigated by immunoblotting and enzyme-linked immunosorbent assay (ELISA). The immunoblotting of hot pepper proteins extracts (HPEs) against serum of hot pepper sensitized patients revealed dominant IgE binding to 14, 37, and 40 kDa molecules. The specific levels of IgE to HPEs sample No. 1, 3, and 7 were much higher than the other samples in patients. Also, IgE binding capacity of HPEs were not reduced by thermal processing and digestion in ELISA using human IgE antibody acquired from hot pepper sensitized patients. By means of Western blotting using anti-thaumatin IgY, thaumatin-like protein (TLP) acting as allergen in several plants and fruits was detected in tested hot peppers. This study demonstrates that the antigenic protein in hot peppers are present but are differently contained according to cultivars.

• 한국어병학회지
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• 제34권1호
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• pp.111-115
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• 2021

Immunoglobulin M (IgM) of sevenband grouper (Epinephelus septemfasciatus) was purified by mannan-binding protein (MBP) affinity column. The purified IgM had an apparent molecular weights of 76 (heavy chain) and 28 (light chain) kDa in sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Eight hybridoma clones secreting monoclonal antibodies (mAbs) against sevenband grouper IgM were established. Antibody detection enzyme-linked immunosorbent assay (ELISA) with bovine serum albumin (BSA, antigen) and the 8 mAbs revealed that optical density (OD) values were clearly different between sera from BSA-immunization and non-immunization of sevenband grouper. These results suggest that the produced mAbs in this study are specifically reacted with IgM of sevenband grouper.

• Asian-Australasian Journal of Animal Sciences
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• 제29권10호
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• pp.1508-1514
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• 2016

A series of antagonists specifically targeting growth hormone receptors (GHR) in different species, such as humans, rats, bovines, and mice, have been designed; however, there are currently no antagonists that target the porcine growth hormone (GH). Therefore, in this study, we developed and characterized a porcine GHR (pGHR) antibody antagonist (denoted by AN98) via the hybridoma technique. The results from enzyme-linked immunosorbent assay, fluorescence activated cell sorter, indirect immunoinfluscent assay, and competitive receptor binding analysis showed that AN98 could specifically recognize pGHR, and further experiments indicated that AN98 could effectively inhibit pGH-induced signalling in CHO-pGHR cells and porcine hepatocytes. In addition, AN98 also inhibited GH-induced insulin-like growth factor-1 (IGF-1) secretion in porcine hepatocytes. In summary, these findings indicated that AN98, as a pGHR-specific antagonist, has potential applications in pGH-pGHR-related research on domestic pigs.

• Asian Pacific Journal of Cancer Prevention
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• 제15권16호
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• pp.6911-6917
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• 2014

Background: Hepatocellular carcinoma is a complex and heterogeneous tumor with poor prognosis due to frequent intrahepatic spread and extrahepatic metastasis. The molecular mechanisms underlying HCC pathogenesis still remain obscure. Objectives: We aimed to investigate the abundance and the DNA binding activity of nuclear factor kappa B/p65 subunit in peripheral blood mononuclear cells from patients with HCC and to assess its prognostic significance and association with hypoxia inducible factor one alpha (HIF-$1{\alpha}$) in blood. Subjects and methods: This study was carried out on 40 patients classified equally into liver cirrhosis (group I) and HCC (group II), in addition to 20 healthy volunteers (group III). All groups were subjected to measurement of NF-${\kappa}B$/P65 subunit expression levels by real time-PCR, and DNA binding activity was evaluated by transcription factor binding immunoassay. Serum HIF-$1{\alpha}$ levels were estimated by enzyme-linked immunosorbent assay (ELISA). Significant increase of both the expression level and DNA binding activity of NF-${\kappa}B$/P65 subunit together with serum HIF-1 alpha levels was noted in HCC patients compared to liver cirrhosis and control subjects, with significant positive correlation with parameters for bad prognosis of HCC. In conclusion, NF-${\kappa}B$ signaling is activated in HCC and associated with disease prognosis and with high circulating levels of HIF-1 alpha.

• Journal of Plant Biotechnology
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• 제36권3호
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• pp.268-274
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• 2009

Porphyromonas gingivalis, the gram-negative anaerobic oral bacterium, initiates periodontal disease by binding to saliva-coated oral surface. The cholera toxin B subunit (CTB) genetically linked to FimA1 (1-200 aa) or FimA2 (201-337 aa) of the P. gingivalis fimbrial antigen were introduced into Solanum tuberosum cells by Agrobacterium tumefaciens-mediated transformation method. The integration of CTB-FimA1 or CTB-FimA2 fusion genes were confirmed in the chromosome of transformed leaves by genomic DNA PCR amplification method. Synthesis and assembly of the CTB-FimA fusion proteins into oligomeric structures with pentamer size was detected in transformed tuber extracts by immunoblot analysis. The binding activities of CTB-FimA fusion proteins to intestinal epithelial cell membrane receptors were confirmed by GM1-ganglioside enzyme-linked immunosorbent assay (GM1-ELISA). The ELISA showed that the expression levels of the CTB-FimA1 or CTB-FimA2 fusion proteins were 0.0019, 0.002% of the total soluble protein in transgenic tuber tissues, respectively The synthesis of CTB-FimA monomers and their assembly into biologically active oligomers in transformed potato tuber tissues demonstrates the feasibility of using edible plants for the production of enterocyte targeted fimbrial antigens that could elicit mucosal immune responses.

• Food Science and Biotechnology
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• 제17권5호
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• pp.919-924
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• 2008

The aim of this study was to observe the changes in allergenicity of saeujeot (salted and fermented shrimp) using a competitive indirect enzyme-linked immunosorbent assay (Ci-ELISA). The fermentation conditions tested for saeujeot consisted of various temperatures (25, 15, and $5^{\circ}C$) and salt concentrations (25, 15, and 10%). When saeujeot was fermented at a low salt concentration and high temperature, the binding ability of mAb and shrimp-allergic patient serum to allergen was significantly decreased. In particular, the binding ability of mAb to allergen in saeujeot fermented with 10% salt at $25^{\circ}C$ for 5 days decreased to 5%. Also, the binding ability of shrimp-allergic patient serum to allergen in saeujeot fermented for 5 days with 10% salt at $25^{\circ}C$ was 8%. In conclusion, the binding of mAb and shrimp-allergic patient serum to tropomyosin in saeujeot decreased with longer fermentation periods, lower salt concentrations (10%), and higher temperatures ($25^{\circ}C$).

• 약학회지
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• 제44권4호
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• pp.340-346
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• 2000

Cervical cancer is one of the leading causes of female death from cancer worldwide with about 500,000 deaths per year. A strong association between certain human papilloma viruses (HPV types 16 and 18) and cervical cancer has been well known. An extract of natural products, named as Somatid, has been used to investigate whether this agent has the ability of inhibiting the oncogenes E6 and E7 of HPV type 16. This Somatid inhibited the proliferation of human cervical cancer cell lines (C-33A, SiHa, CaSki) and HaCaT keratinocytes in a dose response manner, In vitro binding assay and ELISA showed that Somatid inhibited the in vitro biding of E6 and E6AP which are essential for the binding and degradation of the tumor suppressor p53. In addition, Somatid inhibited the in vitro binding of E7 and Rb which is essential tumor suppressor for the control of cell cycle. The levels of mRNA for E6 and E7 were also decreased by Somatid. Our data suggested that Somatid inhibited the oncogenecity of E6 and E7 of HPV 16 type, thus can be used as a putative anti-HPV agent for the treatment of cervical carcinomas caused by HPV.

• Toxicological Research
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• 제27권2호
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• pp.125-131
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• 2011

Through the present study, we produced a monoclonal antibody against aflatoxin B1 (AFB1) using AFB1-carboxymethoxylamine BSA conjugates. One clone showing high binding ability was selected and it was applied to develop a direct competitive ELISA system. The epitope densities of AFB1-CMO against BSA and KLH were about 1 : 6 and 1 : 545, respectively. The monoclonal antibody (mAb) from cloned hybridoma cell was the IgG1 subclass with ${\lambda}$-type light chains. The $IC_{50}s$ of the monoclonal antibody developed for AFB1, AFB2, AFG1 and AFG2 were 4.36, 7.22, 6.61 and 29.41 ng/ml, respectively, based on the AFB1-KLH coated ELISA system and 15.28, 26.62, 32.75 and 56.67 ng/ml, respectively, based on the mAb coated ELISA. Cross-relativities of mAb to AFB1 for AFB2, AFG1 and AFG2 were 60.47, 65.97 and 14.83% in the AFB1-KLH coated ELISA, and 59.41, 46.66 and 26.97% in the mAb coated ELISA, respectively. Quantitative calculations for AFB1 from the AFB1-Ab ELISA and AFB1-Ag ELISA ranged from 0.25 to 25 ng/ml ($R^2$ > 0.99) and from 1 to 100 ng/ml ($R^2$ > 0.99), respectively. The intra- and inter-assay precision CVs were < 10% in both ELISA assay, representing good reproducibility of developed assay. Recoveries ranged from 79.18 to 91.27%, CVs ranged from 3.21 to 7.97% after spiking AFB1 at concentrations ranging from 5 to 50 ng/ml and following by extraction with 70% methanol solution in the Ab-coated ELISA. In conclusion, we produced a group specific mAb against aflatoxins and developed two direct competitive ELISAs for the detection of AFB1 in feeds based on a monoclonal antibody developed.

• Journal of Microbiology
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• 제43권6호
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• pp.537-545
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• 2005

Even though neutralizing antibodies against the Hantaan virus (HTNV) has been proven to be critical against viral infections, the cellular immune responses to HTNV are also assumed to be important for viral clearance. In this report, we have examined the cellular and humoral immune responses against the HTNV nucleocapsid protein (NP) elicited by virus infection or DNA vaccination. To examine the cellular immune response against HTNV NP, we used $H-2K^b$ restricted T-cell epitopes of NP. The NP-specific $CD8^+$ T cell response was analyzed using a $^{51}Cr-release$ assay, intracellular cytokine staining assay, enzyme-linked immunospot assay and tetramer binding assay in C57BL/6 mice infected with HTNV. Using these methods, we found that HTNV infection elicited a strong NP-specific $CD8^+$ T cell response at eight days after infection. We also found that several different methods to check the NP-specific $CD8^+$ T cell response showed a very high correlation among analysis. In the case of DNA vaccination by plasmid encoding nucleocapsid gene, the NP-specific antibody response was elicited $2\~4$ weeks after immunization and maximized at $6\~8$ weeks. NP-specific $CD8^+$ T cell response reached its peak 3 weeks after immunization. In a challenge test with the recombinant vaccinia virus expressing NP (rVV-HTNV-N), the rVV-HTNV-N titers in DNA vaccinated mice were decreased about 100-fold compared to the negative control mice.

• BMB Reports
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• 제36권2호
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• pp.207-213
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• 2003

Peroxisome proliferator-activated receptors (PPARs) are orphan nuclear hormone receptors that are known to control the expression of genes that are involved in lipid homeostasis and energy balance. PPARs activate gene transcription in response to a variety of compounds, including hypolipidemic drugs. Most of these compounds have high affinity to the ligand-binding domain (LBD) of PPARs and cause a conformational change within PPARs. As a result, the receptor is converted to an activated mode that promotes the recruitment fo co-activators such as the steroid receptor co-activator-1 (SRC-1). Based on the activation mechanism of PPARs (the ligand binding to $PPAR{\gamma}$ induces interactions of the receptor with transcriptional co-activators), we performed Western blot and ELISA. These showed that the indomethacin, a $PPAR{\gamma}$ ligand, increased the binding between $PPAR{\gamma}$ and SRC-1 in a ligand dose-dependent manner. These results suggested that the in vitro conformational change of $PPAR{\gamma}$ by ligands was also induced, and increased the levels of the ligand-dependent interaction with SRC-1. Collectively, we developed a novel and useful ELISA system for the mass screening of $PPAR{\gamma}$ ligands. This screening system (based on the interaction between $PPAR{\gamma}$ and SRC-1) may be a promising system in the development of drugs for metabolic disorders.