• BMB Reports
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• 제35권2호
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• pp.206-212
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• 2002

The NS2B-NS3(pro) polyprotein segment from the dengue virus serotype 2 strain 16681 was purified from overexpressing E. coli by metal chelate affinity chromatography and gel filtration. Enzymatic activity of the refolded NS2B-NS3(pro) protease complex was determined in vitro with dansyl-labeled peptide substrates, based upon native dengue virus type 2 cleavage sites. The 12mer substrate peptides and the cleavage products could be separated by reversed-phase HPLC, and were identified by UV and fluorescence detection. All of the peptide substrates (representing the DEN polyprotein junction sequences at the NS2A/NS2B, NS2B/NS3, NS3/NS4A and NS4B/NS5 sites) were cleaved by the recombinant protease NS2B-NS3(pro). No cleavage was observed with an enzymatically inactive S135A mutant of the NS3 protein, or with a modified substrate peptide of the NS3/NS4A polyprotein site that contained a K2093A substitution. Enzymatic activity was dependent on the salt concentration. A 50% decrease of activity was observed in the presence of 0.1M sodium chloride. Our results show that the NS3 protease activity of the refolded NS2B-NS3(pro) protein can be assayed in vitro with high specificity by using cleavage-junction derived peptide substrates.

• Korean Chemical Engineering Research
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• 제50권4호
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• pp.738-742
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• 2012

본 연구에서는 알루미나 나노 템플레이트(anodic alumina oxide; AAO)를 이용하여 신속하면서도 효과적으로 나노입자 및 바이오물질을 분리, 농축할 수 있는 나노필터 소자를 개발하였다. 본 연구에서 사용한 나노필터 소자는 유체의 주입 및 흐름이 가능한 미세유체채널(microfluidic channel)을 PDMS에 패터닝하였다. 위아래로 형성된 PDMS 미세유체채널 사이로, 다양한 크기의 나노 다공을 형성하고 있는 AAO 막을 삽입하여 크기에 따른 나노입자 및 바이오 물질을 분리할 수 있었다. 위아래로 PDMS 유체채널과 AAO 분리막을 집적하고, 최종적으로 아크릴레이트 플락스틱(acrylic plastic)으로 전체 소자를 고정하여 나노필터유체소자를 제작하였다. 완성된 나노필터소자를 이용하여 나노입자의 농축효율 및 은나노입자가 뭉쳐져있는 필터존(filtration zone)으로부터 뎅기 바이러스(dengue virus)를 표면증강라만(surface enhanced Raman scattering)분석법에 의해 검출할 수 있었다.

• 한국생물공학회:학술대회논문집
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• 한국생물공학회 2003년도 생물공학의 동향(XIII)
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• pp.99-99
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• 2003

• Pediatric Infection and Vaccine
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• 제23권1호
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• pp.1-9
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• 2016

The Zika virus, a flavivirus related to dengue and Japanese encephalitis was discovered in the Zika forest in Uganda, 1947. Since Zika virus was first reported in Brazil in May 2015, infections have occurred in at least 40 countries, especially in the Americas. Zika virus infection usually is asymptomatic or causes mild illness, but may be related to severe clinical manifestations, particularly microcephaly and Guillain-$Barr{\acute{e}}$ syndrome. Although the possibility of autochthonous Zika virus transmission in South Korea is low, the imported cases and Zika virus-transmitting mosquito should be adequately monitored and promptly managed. In addition, enhancing preparedness for Zika virus infection are needed.

• Parasites, Hosts and Diseases
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• 제56권1호
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• pp.61-70
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• 2018

We developed a Rapid Diagnostic Test (RDT) kit for detecting IgG/IgM antibodies against Zika virus (ZIKV) using monoclonal antibodies to the envelope (E) and non-structural protein 1 (NS1) of ZIKV. These proteins were produced using baculovirus expression vector with Sf9 cells. Monoclonal antibodies J2G7 to NS1 and J5E1 to E protein were selected and conjugated with colloidal gold to produce the Zika IgG/IgM RDT kit (Zika RDT). Comparisons with ELISA, plaque reduction neutralization test (PRNT), and PCR were done to investigate the analytical sensitivity of Zika RDT, which resulted in 100% identical results. Sensitivity and specificity of Zika RDT in a field test was determined using positive and negative samples from Brazil and Korea. The diagnostic accuracy of Zika RDT was fairly high; sensitivity and specificity for IgG was 99.0 and 99.3%, respectively, while for IgM it was 96.7 and 98.7%, respectively. Cross reaction with dengue virus was evaluated using anti-Dengue Mixed Titer Performance Panel (PVD201), in which the Zika RDT showed cross-reactions with DENV in 16.7% and 5.6% in IgG and IgM, respectively. Cross reactions were not observed with West Nile, yellow fever, and hepatitis C virus infected sera. Zika RDT kit is very simple to use, rapid to assay, and very sensitive, and highly specific. Therefore, it would serve as a choice of method for point-of-care diagnosis and large scale surveys of ZIKV infection under clinical or field conditions worldwide in endemic areas.

• 한국생물공학회:학술대회논문집
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• 한국생물공학회 2005년도 생물공학의 동향(XVII)
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• pp.599-599
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• 2005

• Journal of Plant Biotechnology
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• 제41권1호
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• pp.50-55
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• 2014

Plant-based vaccines possess some advantages over other types of vaccine biotechnology such as safety, low cost of mass vaccination programs, and wider use of vaccines for medicine. This study was undertaken to develop the transgenic maize as edible vaccine candidates for humans. The immature embryos of HiII genotype were inoculated with A. tumefaciens strain C58C1 containing the binary vectors (V662 or V663). The vectors carrying nptII gene as selection marker and scEDIII (V662) or wCTB-scEDIII (V663) target gene, which code EIII proteins inhibite viral adsorption by cells. In total, 721 maize immature embryos were transformed and twenty-two putative transgenic plants were regenerated after 12 weeks selection regime. Of them, two- and six-plants were proved to be integrated with scEDIII and wCTB-scEDIII genes, respectively, by Southern blot analysis. However, only one plant (V662-29-3864) can express the gene of interest confirmed by Northern blot analysis. These results demonstrated that this plant could be used as a candidated source of the vaccine production.

• IMMUNE NETWORK
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• 제16권5호
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• pp.261-270
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• 2016

The human immune system has evolved to fight against foreign pathogens. It plays a central role in the body's defense mechanism. However, the immune memory geared to fight off a previously recognized pathogen, tends to remember an original form of the pathogen when a variant form subsequently invades. This has been termed 'original antigenic sin'. This adverse immunological effect can alter vaccine effectiveness and sometimes cause enhanced pathogenicity or additional inflammatory responses, according to the type of pathogen and the circumstances of infection. Here we aim to give a simplified conceptual understanding of virus infection and original antigenic sin by comparing and contrasting the two examples of recurring infections such as influenza and dengue viruses in humans.

• 한국콘텐츠학회:학술대회논문집
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• 한국콘텐츠학회 2017년도 춘계 종합학술대회 논문집
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• pp.297-298
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• 2017

플라비바이러스(Flavivirus)는 모기와 같은 곤충을 매개로 하여 인체에 감염된다고 잘 알려져 있다. 그 대표적인 예로 지카 바이러스(Zika virus), 뎅기 바이러스(Dengue virus), 황열 바이러스(Yellow fever virus), 일본 뇌염 바이러스(Japanese encephalitis virus) 등을 들 수 있다. 본 연구에서는 생물정보학을 기반으로 인체 감염 주요 플라비바이러스인 지카 바이러스, 뎅기 바이러스. 황열 바이러스, 일본 뇌염 바이러스의 총 4종류 플라비바이러스에 공통적으로 적용 가능한 펩타이드 백신 후보를 제시하고자 한다. 먼저 UniProt (The Universal Protein Resource)의 유전자 서열정보를 이용하여 4종류의 바이러스가 가진 단백질 중 백신으로써 적합한 단백질을 선정하였다. 선정된 단백질의 아미노산 서열정보를 바탕으로 IEDB (Immune Epitope Database And Analysis Resource)를 활용한 에피토프(epitope) 분석을 통해 에피토프로 작용하는 4 종류 바이러스의 공통적인 서열을 도출하였다.

• IMMUNE NETWORK
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• 제7권2호
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• pp.66-74
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• 2007

Background: The genus Flavivirus consists of many emerging arboviruses, including Dengue virus (DV), Japanese encephalitis virus (JEV) and West Nile virus (WNV). Effective preventive vaccines remain elusive for these diseases. Mice are being increasingly used as the animal model for vaccine studies. However, the pathogenic mechanisms of these viruses are not clearly understood. Here, we investigated the interaction of DV and JEV with murine bone marrow-derived dendritic cells (bmDC). Methods: ELISA and FACS analysis were employed to investigate cytokine production and phenotypic changes of DCs obtained from bone marrow following flavivirus infection. Results: We observed that these viruses altered the cytokine profile and phenotypic markers. Although both viruses belong to the same family, JEV-infected bmDC produced anti-inflammatory cytokine (IL-10) along with pro-inflammatory cytokines, whereas DV infection induced production of large amounts of pro-inflammatory cytokines (IL-6 and TNF-${\alpha}$) and no IL-10 from murine bmDCs. Both flaviviruses also up-regulated the expression of co-stimulatory molecules such as CD40, CD80 and CD86. JEV infection led to down-regulation of MHC II expression on infected bmDCs. We also found that cytokine production induced by JEV and DV is MyD88-dependent. This dependence was complete for DV, as cytokine production was completely abolished in the absence of MyD88. With regard to JEV, the absence of MyD88 led to a partial reduction in cytokine levels. Conclusion: Here, we demonstrate that MyD88 plays an important role in the pathogenesis of flaviviruses. Our study provides insight into the pathogenesis of JEV and DV in the murine model.