• Journal of fish pathology
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• v.27 no.1
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• pp.1-9
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• 2014

The olive flounder, Paralichthys olivaceus is susceptible to viral hemorrhagic septicaemia virus (VHSV) at $15^{\circ}C$ but no mortality at $20^{\circ}C$ even though the virus can grow well in vitro at $20^{\circ}C$. Thus, we designed an experiment to know immune response of olive flounder against VHSV when the host reared at $15^{\circ}C$ or $20^{\circ}C$. cDNA microarray analysis was performed to compare the gene expression patterns of the kidney cells between the host reared at $15^{\circ}C$ or $20^{\circ}C$. The expression of MHC class I, IL-8, myeloperoxidae and endonuclease G-like having function for the antigen presentation and chemokine-factor were up-regulted both the $15^{\circ}C$ and $20^{\circ}C$ during VHSV infection. MHC class II gene existing on antigen-presenting cells and B cell lymphocytes, immunoglobulin (Ig) genes and phagocytosis related genes were down-regulated at $15^{\circ}C$ but highly expressed at $20^{\circ}C$. It can be thought that innate immune related antigen presentation by MHC class I and phagocytosis reaction against VHSV are efficiently occur both the temperature but macrophage or B cell related antigen presentation via MHC class II fails to induce downstream immune reactions (adaptive immunity) to make antibody, and it can be one of the reason that causes high mortality only at $15^{\circ}C$.

• Journal of the korean academy of Pediatric Dentistry
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• v.33 no.3
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• pp.401-410
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• 2006

Dental caries results from localized demineralization of tooth enamel by acids of bacterial origin produced from the fermentation of dietary sugars. A group of related oral bacteria, collectively known as mutans streptococci, are implicated as the primary etiological agents of human caries. Within this group, Streptococcus mutans has been known as a causative agent for dental caries. As well as acid production yielding the demineralization of tooth enamel, adherence and colonization of S. mutans to the teeth are also important for their virulence Cell-surface fibrillar proteins, which mediate adherence to the salivary pellicle are virulence components of mutans streptococci, and primary candidates for a human caries vaccine. Here we report that the AgI/II gene from S. mutans GS-5 were cloned by PCR amplification of the bacterial chromosomal DNA and the integrity of cloned genes were confirmed by nucleotide sequencing. Sequence analyses showed the sequence alignment of 280 nucleotides between the cloned AgI/II and the reported sequence of S. mutans GS-5 showed the perfect match The cloned genes which signal nucleotide was truncated, were transferred into bacterial expression vector and the recombinant proteins were purified as His-tag fusion proteins In order to generate polyclonal antibodies against the recombinant proteins, AgI/II mr, some $100{\mu}g$ of the proteins was injected into mice three times. It can be used for an effective vaccine production to prevent dental caries caused by pathogenic S. mutans.

• Proceedings of the Korean Vacuum Society Conference
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• 2014.02a
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• pp.436-436
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• 2014

본 연구에서는 전 세계적으로 활발히 연구되고 있는 나노바이오센서 분야 중 가장 주목을 받고 있는 LSPR 원리를 이용한 바이오센서를 제작하였다. 금속 나노입자의 국소 표면 플라즈몬 공명현상에 의한 주위환경에 민감하게 반응하는 특성은 고감도 광학형 바이오센서, 화학물질 검출 센서등에 응용된다. 특히 금 나노막대와 같은 1차 나노구조물은 나노막대의 주변 환경 변화에 따라 뚜렷한 플라즈몬 흡수 밴드 변화를 나타냄으로 센서로 적용 했을 때 고감도의 측정이 가능하다. 본 연구에서는 다공성인 알루미늄 양극산화 박막 주형틀을 이용하여 다양한 종횡비를 가지는 금 나노막대를 합성하고, 나노막대 어레이 형태의 박막을 제작하였다. 금 나노막대의 합성은 알루미늄 양극산화막을 사용한 주형제조 방법(template method)을 사용하는 전기화학 증착법을 사용하였다. 우선 부도체인 알루미늄 양극 산화막의 한쪽면을 열증착 장비를 사용하여 금을 증착하여 작업 전극(working electrode)을 형성하였다. 백금 선(platinum wire)을 보조 전극(counter electrode)으로 사용하고 Ag/AgCl 전극을 기준 전극(reference electrode)으로 사용하여 삼전극계(three-electrode system)를 형성하였으며, 금 도금 용액(orotemp 24 gold plating solution, TECHNIC INC.)을 사용하여, 800 mV 전압에서 금 나노 막대를 합성하였다. 금 나노막대의 길이는 테플론 챔버를 통과한 전하량 또는 전기 증착 시간에 비례하여 결정된다. 금 나노막대를 성장시킨 알루미늄 양극산화막을 실리콘 웨이퍼에 은 페이스트를 사용하여 고정시킨 후 수산화나트륨 (NaOH)용액을 사용하여 알루미늄 양극산화막을 녹여내어 수직방향으로 정렬되어 있는 나노 막대 어레이 박막을 제조 하였다. 또한 제작된 금 나노막대 어레이의 광학적 특성을 평가하였다. 본 연구에서와 같이 나노막대를 직경방향으로 측정할 경우, 직경방향의 transverse mode만 측정된다. 금 나노 막대가 알루미늄 양극산화막 안에 포함된 상태로 측정된 금 나노로드 어레이 박막의 광 스펙트럼 분포는 금 나노막대의 가시광영역에서의 흡수 스펙트럼을 측정하였을시 직경 및 길이에 따라 transverse mode의 ${\lambda}$ max (최대 흡광)의 위치가 변화됨을 나타낸다. 실험 결과를 바탕으로 나노막대의 종횡비가 증가함에 따라 흡수 스펙트럼의 transverse mode ${\lambda}$ max가 미약하게 단파장 영역으로 이동하는 것을 확인할 수 있다. 이러한 결과는 원기둥 형태의 금 나노막대의 흡수 스펙트럼에 대한 이론적인 예측과 부합한다. 바이오센서로의 적용 가능성을 확인하기 위하여 자기조립단분자막을 형성하여 항체를 고정하고 CRP에 대한 응답특성을 평가하였다. CRP 항원-항체의 면역반응에 대한 실험 결과 CRP 항원의 농도가 증가함에 따라 넓은 측정범위에서 선형적으로 흡광도가 증가하는 결과를 나타내었으며, CRP 10 fg/ml의 농도까지 검출할 수 있었다. 센서의 선택성을 확인하기 위하여 감지하고자하는 대상물질이 아닌 Tn T 항원을 감지막에 반응시켜 흡광도 변화를 분석하였다. 결과적으로 제작된 센서칩은 선택성을 가지고 측정하고자하는 물질에만 반응함을 확인하였다. 이러한 결과는 다양한 직경을 사용한 부가적인 LSPR현상의 연구에 활용될 수 있을 것이다.

• Korean Journal of Clinical Laboratory Science
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• v.49 no.3
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• pp.220-226
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• 2017

Dopamine (DA) and serotonin (5-Hydroxytryptamine, 5-HT) are neurotransmitters and hormones that exist in small amounts but have important role in the body. Serum and 24-hour urine are used as specimens, and are usually examined by HPLC-MS. In this study, we tried to detect DA and 5-HT by competitive ELISA using antigen-antibody (Ab) reaction. After immobilizing $5{\mu}g/mL$ BSA conjugate on a 96-well surface, hormone and primary Ab, which are respectively diluted to different concentrations, were treated. Then, HRP-conjugated secondary Ab and TMB were added to measure absorbance. The regression equation and $R^2$ value were calculated based on absorbance, and sensitivity of Ab to hormone as well as the correlation between hormone concentration and absorbance were determined. In DA ELISA, $R^2$, the correlation between the concentration of hormone and absorbance, was the highest by 0.91 when anti-dopamine Ab was diluted 6,000 times and 7,000 times. In 5-HT ELISA, $R^2$ was bigger than 0.90 in every concentration except 3,000 times and 6,000 times. Both DA and 5-HT were not effectively detected at low concentrations (less than $1.0{\times}10^{-7}M$); and because reference value of serum DA is lower than this, HPLC-MS was required to detect serum DA. However, competitive ELISA may be effective in detecting 24-hour urine DA, serum, and 24-hour 5-HT. Further studies are needed to detect hormones more accurately at lower concentrations.

• Clinical and Experimental Reproductive Medicine
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• v.33 no.3
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• pp.171-178
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• 2006

Objective: Human embryonic stem (ES) cells have a great potential in regenerative medicine and tissue engineering. The human ES cells could be differentiated into specific cell types by treatments of growth factors and alterations of gene expressions. However, the efficacy of guided differentiation and isolation of specific cells are still low. In this study, we characterized isolated cells from differentiated human ES cells by magnetic activated cell sorting (MACS) system using specific antibodies to cell surface markers. Methods: The undifferentiated hES cells (Miz-hESC4) were sub-cultured by mechanical isolation of colonies and embryoid bodies were spontaneously differentiated with DMEM containing 10% FBS for 2 weeks. The differentiated cells were isolated to positive and negative cells with MACS system using CD34, human epithelial antigen (HEA) and human fibroblast (HFB) antibodies, respectively. Observation of morphological changes and analysis of marker genes expression were performed during further culture of MACS isolated cells for 4 weeks. Results: Morphology of the CD34 positive cells was firstly round, and then it was changed to small polygonal shape after further culture. The HEA positive cells showed large polygonal, and the HFB positive spindle shape. In RT-PCR analysis of marker genes, the CD34 and HFB positive cells expressed endodermal and mesodermal genes, and HEA positive cells expressed ectodermal genes such as NESTIN and NF68KD. The marker genes expression pattern of CD34 positive cells changed during the extension of culture time. Conclusion: Our results showed the possibility of successful isolation of specific cells by MACS system from undirected differentiated human ES cells. Thus, MACS system and marker antibodies for specific cell types might be useful for guided differentiation and isolation of specific cells from human ES cells.

• Journal of Periodontal and Implant Science
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• v.36 no.3
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• pp.579-590
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• 2006

1. 연구배경 교원질 분해작용을 하는 호중구의 세포질 효소인 기질금속단백분해효소-8은 치주질환, 류마티스 관절염, 그리고 궤양결장염과 같은 염증성 질환에서 농도가 증가한다고 알려져 있다. 최근에는 A. actinomycetemcomitans의 leukotoxin이 사람호중구에서 기질금속단백분해효소-8의 분비를 유도하는 것이 보고되었다. 이 연구의 목적은 선천면역 체계에서 세포표면 항원무리14, Toll-like 수용기, 그리고 $NF-{\kappa}$ B경로를 통하여 A. actinomycetemcomitans의 지질다당질로 유도된 기질금속단백분해효소-8의 분비 여부와 세포기전을 알아보고자 하였다. 2. 연구재료 및 방법 건강한 개인 제공자(남자 13명, 여자 3명)로부터 얻은 개개인의 20ml 말초혈액을 제조사의 지침에 따라 호중구를 추출한 후 항세포표면 항원무리14와 함께 $4^{\circ}C$에서 30분간 전배양 한 후, $37^{\circ}C$에서 9시간 동안 배양시켰다. 추출한 호중구에 Toll-like 수용기 억제제 또는 $NF-{\kappa}$ B억제제인 TPCK를 첨가한 후 $37^{\circ}C$에서 1시간 동안 전배양하고 $37^{\circ}C$에서 9시간 동안 배양시켰다. 호중구에 세포뼈대 억제제인 cholchicine, nocodazole, demecolcine, 그리고 cytochalasin B를 A. actinomycetemcomitans의 지질다당질과 함께 $37^{\circ}C$에서 9시간 동안 배양시켰다. 기질금속단백분해효소-8 분비량은 효소면역측정법을 통해 결정하였다. 통계처리는 일원배치 분산분석법을 이용하였다(p<0.05). 3. 결과 A. actinomycetemcomitans 지질다당질은 기질금속단백분해효소-8의 분비를 증가시켰다. 기질금속단백분해효소-8의 분비는 항세포표면 항원무리14에 의해서 억제되었지만, 항 Toll-like 수용기2, 항 Toll-like 수용기4 항체는 억제시키지 못했다. $NF-{\kappa}$ B 억제제는 A. actinomycetemcomitans의 지질다당질로 유도된 $NF-{\kappa}$ B 결합 활성도와 기질금속단백분해효소-8 분비를 억제하였다. 미세섬유 중합반응 억제제는 A. actinomycetemcomitans의 지질다당질로 유도된 기질금속단백분해효소-8의 분비를 억제시켰으나, 미세관 중합반응억제제는 억제시키지 못했다. 4. 결론 위의 연구결과를 종합하여 볼 때, 기질금속단백분해효소-8은 A. actinomycetemcomitans의 지질다당질로 유도되며, 세포표면 항원무리-$NF-{\kappa}$ B 경로를 통하여 분비되고, 이 분비 과정은 미세섬유 계통이 관여하는 것으로 보인다.

• Journal of Preventive Medicine and Public Health
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• v.33 no.2
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• pp.226-230
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• 2000

Objectives : The purpose of this study was to suggest a proper method for the detection of heaptitis B surface antibody(anti-HBs) in a screening program for hepatitis B vaccination. Methods : Sensivitity, specificity and predictive values were compared between Immunochromatographic assay (ICA) and passive hemagglutination(PHA) in 978 subjects(565 males, 413 females, 19-78 years ranging in age, mean 46.5 years old). EIA was used as a standard method for the detection of HBsAb. Results : Sensitivity in the detection of anti-HBs of PHA and ICA was 88.7%, and 94.9%, specificity was 94.3% and 96.6%, negative predictive value was 96.5%, and 98.0%, and positive predictive value was 82.3%, and 91.3%,, respectively. False negative rate(11.3%) of PHA was higher than that(5.1%) of ICA. The higher the titer of anti-HBs in EIA was, the lower the false negative rate was. There was no false negative result in the cases with $101mIU/{\beta}c$ or more in EIA Conclusion : We suggest that ICA should be the choice of screening method in the detection of anti-HBs in Hepatitis B vaccination program.

• Journal of Pharmacopuncture
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• v.11 no.1
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• pp.5-12
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• 2008

Objectives : Showing that Bong-Han corpuscles(BHC) are suppliers of the stem cells in adulthood, and the Bong-Han ducts(BHD) are transportation routes of stem cells. Methods : BHC and BHD were obtained from the internal organ-surfaces of rats. The sliced BHC and BHD were immunostained with various stem cell markers. Extracellular matrices were also analyzed by immunohistochemistry. Result : The presence of mesenchymal stem cells was confirmed by the expression of Integrin beta 1, Collagen type 1 and Fibronectin. But CD54 was not expressed. The hematopoietic stem cell marker, Thy 1 was strongly expressed. BHDs showed Collagen type 1, Fibronectin, and vWF expression. Conclusion : Both hematopoietic and mesenchymal stem cell markers were expressed strongly in BHC similarly as in bone marrow. An endothelial cell marker(vWF) demonstrated the possibility of the stem cell transportation routes of BHD.

• Proceedings of the KIEE Conference
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• 2004.07c
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• pp.1579-1581
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• 2004

바이오센서는 효소(enzyme), 생분자(biomolecule), 항체(antibody), 세포(cell) 등의 biological agent를 인지 물질(recognition material)로 하여 측정하고자하는 분석 대상(analyte)과 높은 선택성으로 반응을 일으키게 하여 그 결과를 기존의 물리, 화학센서로 감지 해내는 방식이므로 기존의 의료용 화학센서를 대체하는 추세이다. 바이오센서가 기존의 센서와 구별되는 점은 생물질의 선택적인 반응 및 결합을 이용하는 것이므로 바이오센서의 실용화에 있어서 가장 중요한 것은 생체 반응 물질의 고정화 기술과 고정화막의 선택이라 할 수 있다. 일정전압법을 이용한 요소센서는 많이 연구되어 오고 있으나 낮은 농도에서의 감도저하에 따른 단점으로 상용화에 이르지 못하고 있다. 본 논문은 요소센서의 이용하기 위한 고정화막으로 3-mercaptopropionic acid 자기조립 단인층의 전기화학적 특성을 고창하였다. 자기조립 단일층은 직접적인 전자전달로 인하여 낮은 요소 농도에서 뛰어난 강도와 빠른 반응 시간을 보였으며, 특히 다공질 실라콘을 기질로 사용한 경우 평면 전극 보다 약 3배의 감도 증가 효과를 가져왔다. 자기조립 단일층의 표면 분석은 X-ray photoelectron spectroscopy(XPS)를 이용하였다.

• 한국생물공학회:학술대회논문집
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• 2001.11a
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• pp.231-234
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• 2001

We performed a basic experiment for rapid. on-line, real-time measurement of HBsAg by using a BIACORE biosensor, a chip-based sensor utilizing surface plasmon resonance technology to quantify the recognition and interaction of biomolecules. We immobilized an a -HBsAg antibody on a CM5 chip surface which was activated by N-hydroxysuccinimide for amine coupling with HBsAg, and measured the mass increase from the coupling. This study showed the potential of this biosensor-based method as a rapid, multi-sample, on-line assay. Once properly validated, it can serve as a more powerful method for HBsAg quantification.