• 대한방사성의약품학회지
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• 제3권2호
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• pp.54-58
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• 2017

Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry(MALDI-TOF MS) is one of the powerful methods that enable analysis of small molecules as well as large molecules up to about 500,000 Da without severe fragmentation. MALDI-TOF MS, thus, has been a very useful an analytical tool for the confirmation of synthetic molecules, probing PTMs, and identifying structures of a given protein. In recent nuclear medicine, MALDI-TOF MS liner ion mode helps researcher calculate the average number of chelator(or linkage) per an antibody conjugate, such as DOTA-(or DFO-) trastuzumab for labeling a medical radioisotope. This simple technique can be utilized to improve the labeling method and control the quality at the development of antibody-based radiopharmaceuticals, which is very effected to diagnosis and therapy for in vivo tumor cells, with radioisotopes like $^{89}Zr$, $^{64}Cu$, and 177Lu. To minimize the error, MALDI-TOF MS measurement is repeatedly performed for each sample in this study, and external calibration is carried out after data collection.

• Clinical and Experimental Reproductive Medicine
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• 제44권1호
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• pp.15-21
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• 2017

Objective: The aims of this study were to investigate whether fertilization could induce the resumption of meiosis in mouse oocytes arrested at metaphase I (MI) after in vitro maturation (IVM), and to investigate the effect of $Ca^{2+}$ chelator treatment at the time of fertilization on the transition from MI to metaphase II (MII). Methods: MII-stage and arrested MI-stage mouse oocytes after IVM were fertilized, and then embryonic development was monitored. Blastocysts from each group were transferred into 2.5 days post-coitum pseudo-pregnant ICR mice. MI oocytes after IVM were treated with a $Ca^{2+}$ chelator to investigate the effect of $Ca^{2+}$ oscillations on their maturation. Results: As insemination time increased, the number of oocytes in the MI group that reached the MII stage also increased. The blastocyst rates and total cell numbers in the MII group were significantly higher than in the MI group. No pregnancy occurred in the MI group, but 10 pregnancies were achieved (10 of 12) in the MII group. The proportion of MI oocytes that matured to MII oocytes after fertilization was significantly higher in the non-treated group than in the $Ca^{2+}$ chelator-treated group. Conclusion: The findings that a higher proportion of MI-arrested oocytes progressed to MII after fertilization and that the MI-to-MII transition was blocked by $Ca^{2+}$ chelator treatments before fertilization indicate that the maturation of MI oocytes to MII oocytes is associated with intracellular $Ca^{2+}$ oscillations driven by fertilization.

• Journal of Plant Biotechnology
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• 제34권3호
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• pp.213-221
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• 2007

분리된 무 자엽 조직에 산화질소 (nitric oxide: NO) 공여체인 sodium nitroprusside (SNP) 처리 시 농도 의존방식으로 부정근의 발달을 증진시켰다. 그러나 이러한 NO 증진 효과는 세포외 칼슘 chelator인 0.5 mM EGTA 또는 세포막 칼슘채널 차단제인 0.1 mM $LaCl_3$를 각각 $50\;{\mu}M$ SNP와 함께 혼합처리 시 반전되었다. 또한, 뿌리 발생에서 중심적 역할을 수행하는 것으로 알려진 guaiacol peroxidase (GPX)와 syringaldazine peroxidase (SPX)의 활성도가 SNP 단독 처리된 자엽에서 부정근이 형성되는 동안 현저히 증가하였다. 그러나, SNP와 $LaCl_3$ 혼합처리 시 SNP에 의해 유도된 GPX와 SPX 활성도 증가가 거의 증류수 대조구 수준으로 억제되었다. calmodulin의 anatagonist인 trifuoperazine 역시 SNP로 처리된 자엽에서 부정근 형성을 억제하여 발생된 뿌리의 개수와 길이를 감소시켰으며 동시에 GPX와 SPX를 불활성화 하였다. 결론적으로, 이들 결과는 칼슘이 GPX와 SPX 활성도 조절을 통해 부정근 유도를 이끄는 NO 반응에 포함되어 있음을 나타내는 것이다.

• Nuclear Medicine and Molecular Imaging
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• 제43권5호
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• pp.487-494
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• 2009

목적: 작은 크기의 재조합 단일사슬 항체는 빠른 혈중 제거율과 종양의 항체 집적율이 증가되는 등의 장점을 가지고 있다. 반면에 항체의 작은 크기는 방사성 또는 형광물질의 표지를 위한 킬레이터 결합에 중요한 아미노산 그룹의 감소를 의미하기도 한다. 본 연구에서는 단일사슬 lym-1 염기서열 C-말단에 lysine 아미노산 태그를 삽입하여 형광 물질의 직접표지 및 그 표지수율 증가를 확인하고자 하였다. 대상 및 방법: 대장균 pET-22b (+) 벡터에 재조합 된 lysine 삽입 단일사슬 lym-1유전자는 대장균 BL21 (DE3)에 형질전환하여 발현하였다. 생산된 lysine lym-1 항체는 Ni-NTA 컬럽과 분자량 컬럼을 사용해 정제하였고. 단백질 전기 영동과 western blot을 통해 확인하였다. lysine lym-1 항체에 방사성 동위원소인 I-124, I-125, I-131 과 Tc-99m를 표지하여 그 수율을 확인하였으며 유세포계측기를 사용해 형광물질인 FITC가 직접표지된 라이신 lym-1 항체의 면역반응성을 사람의 버킷 림프종 세포주인 Raji 세포주에서 면역반응성을 확인하였다. 결과 Lysine도입 단일사슬 lym-1 항체는 두 과정의 정제를 통하여 획득하였으며 그 크기는 약 48 KDa이었고, 방사성동위원소인 I-124, I-125, I-131과 Tc-99m의 표지수율은 각각 >99%, >99%, >95%, >99%로 확인되었다. 유세포계측을 통한 lysine 도입 단일사슬 lym-1항체의 면역반응성은 기존의 단일사슬 lym-1항체와 유사함을 확인하였다. 결론: 재조합 lym-1 항체에 형광 물질을 직접 표지하기 위한 lysine 아미노산의 도입은 항체의 면역반응성 감소를 최소화 시키면서 직접표지 수율을 증가시킬 수 있는 유용한 방법임을 확인하였다.

• 대한방사성의약품학회지
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• 제1권1호
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• pp.1-8
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• 2015

It is essential use of long term half life radioisotopes for positron emission tomography (PET) imaging study of biopharmaceuticals because most of biopharmaceuticals have long biological half-life. Some representative isotopes are $^{124}I$, $^{64}Cu$, $^{89}Zr$ and so on. These PET radioisotopes and their radiopharmaceuticals have recently received growing interest because of long half life and good imaging properties. Furthermore, $^{64}Cu$ and $^{89}Zr$ can be used in a number of radiopharmaceuticals due to its ease of conjugation to peptides and antibodies using the proper chelator. In recent years, since $^{124}I$ was first developed in 2005, we have been studied to develop an efficient method and procedure for producing these radioisotopes, and we have made considerable progress in production of long term half life radioisotopes. This review introduces the general production system, purification procedure, and several advances on targeting method for $^{124}I$ and $^{64}Cu$ in KIRAMS.

• 한방재활의학과학회지
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• 제20권1호
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• pp.27-36
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• 2010

목 적 : 본 연구는 전침 자극에 의한 척수내 N-methyl-D-aspartate(NMDA) 수용체의 NR1 및 NR2B subunit 인산화에 미치는 세포내 칼슘 저해제 bis-(2-aminophenoxy)-ethane-N,N,N',N'-tetraaceticacid(BAPTA)의 영향을 조사하였다. 방 법 : 인체의 족삼리(足三里)(ST36)와 삼음교(三陰交)(SP6)에 해당하는 혈자리에 2 Hz전침 자극을 1.0mA 세기로 30분 동안 자극하였다. 결 과 : 전침 무통각을 측정한 결과 높은 농도의 BAPTA 복강내주사 처리군에서 저하가 관찰되었다. 전침 처치 후 60분 후 분리한 $L_{4-5}$ 척수 분절에서 C-fos 발현 신경세포 수는 BAPTA 처리에 의해 감소하는데 특히 고유핵에서 저하가 현저하였다. 평균 integrated optical density로 비교한 NR1과 NR2B subunit에 대한 면역조직화학적 발현을 보면 전침 자극은 정상군에 비해 얕은 층판에서 증가하였다. NR1과 NR2B subunit의 인산화형에 대한 발현을 보면 NR1 인산화형은 척수 배각 전 부위에서, NR2B 인산화형은 얕은 층판에서 증가하였으며 BAPTA 처리에 의해 NR1 인산화형은 얕은 층판과 목 부위에서 NR2B 인산화형은 얕은 층판에서 현저한 감소를 보였다. western blot로 살펴본 BAPTA 처리에 의한 NR1 및 NR2B 인산화형 변화는 면역조직화학적 방법과 유사한 결과를 보여 주었다. 결 론 : 전침 무통각은 세포내 칼슘에 의한 척수 내 NMDA 수용체의 인산화가 중요한 요인으로 작용할 가능성이 있다.

• Toxicological Research
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• 제22권1호
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• pp.47-54
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• 2006

One of the main attractions of treatment with herbal medicine is its apparent lack of side effects compared with the drug therapies used in allopathic medicine. However, evidence from various countries suggest that Asian herbal medicine carry a significant risk of contamination with toxic heavy metals at levels that may seriously threaten health. The aims of this study were to analyze and compare concentrations of heavy metals in urine and hair from 184 patients taking herbal medicines in the form of decoctions and/or pills in comparison to 101 control subjects taking either Western or no medications. Levels of metal concentrations exceeding WHO reference values were observed in a number of hair and urine samples for all subjects. After adjusting for potential confounders, taking decoctions or pills was associated with higher levels of some metals (such as Cu, Pb in urine), as well a higher odds ratio of exceeding the upper limit of reference ranges for Pb, Hg in hair. In contrast, taking decoctions or pills was associated with lower levels of some metals (such as Cu in urine and Cd, Cu, Hg, Pb in hair), suggesting that some herbal medicines may have a chelating effect on heavy metals in the body. Overall, the results obtained in the study show a mixed picture and suggest that heavy metals contamination in herbs is sometimes present, but may also be counteracted by the potential for some herbal medicines to act as chelating agents. Further study must be followed to obtain more concrete evidence.

• Journal of Applied Biological Chemistry
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• 제43권1호
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• pp.5-11
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• 2000

Three cDNA clones encoding rice calmodulin (CaM) isoforms (OsCaM-1, OsCaM-2, and OsCaM-3) were isolated from a rice cDNA library constructed from suspension-cultured rice cells treated with fungal elicitor. The coding regions of OsCaM-1 and O.sCaM-2 were 89% homologous at DNA Ievel, whereas the 5' and 3' untranslated regions were highly divergent. The polypeptides encoded by OsCaM-1 and OsCaM-2 was identical except two conservative substitution at position 8 and 75. The coding region of OsCaM-3 was consist of a typical conserved CaM domain and an additional C-terminal extension. The amino acid sequence of conserved CaM domain of OsCaM-3 shared only 86% identity with that OsCaM-1. The OsCaM-3 cDNA is belongs to a novel group of calmodulin gene due to its C-terminal extension of 38 amino acids, a large number of which are positively charged. The extension also contains a C-terminal CaaX-box prenylation site (CVlL). Genomic Southern analysis revealed at least six copies of CaM or CaM-related genes, suggesting that calmodulin may be represented by a small multigene family in the rice geneme. Expression of OsCaM gene was examined through Northern blot analysis. Transcript level of OsCaM-3 was increased by treatment with a fungal elicitor, whereas the OsCaM-1 and OsCaM-2 genes did not respond to the fungal elicitor. The expression of OsCaM-3 gene was remarkable inhibited in the rice cells treated with cyclosporine A, calcinurin inhibitor.

• Archives of Pharmacal Research
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• 제27권9호
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• pp.961-967
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• 2004

The development of an antibody labeling method with $^{99m}$Tc is important for cancer imaging. Most bifunctional chelate methods for $^{99m}$Tc labeling of antibody incorporate a $^{99m}$Tc chelator through a linkage to lysine residue. In the present study, a novel site-specific $^{99m}$Tc labeling method at carbohydrate side chain in the Fc region of 2 antibodies (T101 and rabbit anti-human serum albumin antibody (RPAb)) using dihydrazinophthalazine (DHZ) which has 2 hydrazino groups was developed. The antibodies were oxidized with sodium periodate to pro-duce aldehyde on the Fc region. Then, one hydrazine group of DHZ was conjugated with an aldehyde group of antibody through the formation of a hydrazone. The other hydrazine group was used for labeling with $^{99m}$Tc. The number of conjugated DHZ was 1.7 per antibody. $^{99m}$Tc labeling efficiency was 46-85％ for T101 and 67∼87％ for RPAb. Indirect labeling with DHZ conjugated antibodies showed higher stability than direct labeling with reduced antibodies. High immunoreactivities were conserved for both indirectly and directly labeled antibodies. A biodistribution study found high blood activity related to directly labeled T1 01 at early time point as well as low liver activity due to indirectly labeled T101 at later time point. However, these findings do not affect practical use. No significantly different biodistribution was observed in the other organs. The research concluded that DHZ can be used as a site-specific bifunctional chelating agent for labeling antibody with $^{99m}$Tc. Moreover, $^{99m}$Tc labeled antibody via DHZ was found to have excellent chemical and biological properties for nuclear medicine imaging.edicine imaging.

• Applied Microscopy
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• 제38권1호
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• pp.1-10
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• 2008

본 연구는 급성 수은 독성에 대한 민들레의 효과를 알아보기 위하여 시도되었다. 30 g 내외의 생쥐를 대상으로 대조군, 수은(5 mg/kg) 투여군, 수은 투여 후 민들레(3 g/kg)를 구강투여 한 후 다시 24, 48, 72, 96시간, 1주일군으로 세분하여 간장 손상 억제효과를 알아보기 위하여 생화학적 및 조직학적 실험을 실시하였다. 혈액중 aspartate amiotransferase (AST)와 alanine aminotransferase (ALT)의 수치는 민들레 투여군이 수은 투여군보다 감소되었다. 또한 간조직의 SOD와 catalase활성도 역시 민들레 투여군이 수은 투여군에 비하여 감소하였으나 통계학적 유의성은 없었다. 간 조직의 광학현미경적 관찰에서 심한 조직괴사가 관찰되지만 민들레 투여군에서는 문맥주위의 약간의 괴사와 심한 호중구 침윤현상이 관찰되었다. 전자현미경적 관찰 결과, 간장에서 수은 투여군은 간세포의 핵이 함입되어 불규칙했으며 미토콘드리아와 조면소포체의 수조가 팽대되고, 리보솜의 탈락이 관찰되었다. 민들레 투여군은 핵이 정상적인 상태로 관찰되었고, 전자밀도가 높은 미토콘드리아가 분포되어 있었으며, 리보솜이 부착된 상태로 층판구조를 형성하는 조면소포체가 관찰되었다. 이상의 연구 결과로 보아 민들레가 수은으로 유발된 간손상을 보호하는 효과가 있는 것으로 사료되며 민들레에 대한 다양한 연구가 필요할 것으로 사료된다.