• KSBB Journal
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• 제5권4호
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• pp.315-321
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• 1990

Rabbit anti-bovine heart PDH antiserum was raised against El(a, b) isolated from PDC, and then applied to detect Ela and Elb. Appropriate amounis of El were fractionated by SDS-PAGE and electrophoretically transferred to nitrocellulose membrane. The Ela and Elb on the membrane were incubated with anti-El antiserum and identified by GAR-HRP system. It has been found that the immunodetection sensitivity of Ela and Elb were directly proportional to the amount of antigen and transfer time. The lengthy transfer times increased the immunodetection sensitivity of Ela and Elb. The maximal detection sensitivity of Western blotting of Ela and Elb was achieved at 3.5 V/cm for 16-hour transfer under these experimental conditions.

• Applied Biological Chemistry
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• 제43권4호
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• pp.231-235
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• 2000

담배 배양 세포의 성장과정 중의 칼모듈린 농도변화 및 칼모듈린 결합 단백질의 종류에 대하여 조사하고 이들 단백질들 중 글루탐산 탈탄산효소를 immunodetection과 활성측정으로 확인하였다. 담배세포는 유도기(초기 $1{\sim}2$일간), 대수증식기($3{\sim}5$일), 정지기 등의 전형적인 성장 패턴을 보였다. 칼모듈린의 농도는 비록 대수증식기에 약간 감소하는 경향을 보이다 정지기에 이르면서 유도기의 수준을 회복하는 것으로 나타났지만 전체적으로는 성장단계에 관계없이 유사한 수준을 유지하는 것으로 나타났다. 주요 칼슘-의존형 칼모듈린 결합단백질은 56, 46, 36, 32-kDa의 4종류인 것으로 조사되었고, 모노클로날 항체를 이용하여 immunodetection을 실시해 본 결과 56-kDa 단백질이 담배 글루탐산 탈탄산효소로 확인되었다. 56-kDa의 글루탐산 탈탄산효소는 대수증식기에 수확한 세포에서 가장 많이 검출되었고, 이와같은 패턴은 효소활성 측정에서도 확인되었다. 이러한 결과들은 담배세포의 성장과정 중에 칼슘/칼모듈린-의존형 글루탐산 탈탄산효소 농도가 조절되고 있음을 제안해 주는 것이다.

• Journal of Microbiology and Biotechnology
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• 제28권7호
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• pp.1185-1198
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• 2018

The nucleosomal organization of chromatin using histone proteins is a fundamental and ubiquitous feature of eukaryotic nuclei, with the major exception of dinoflagellates. Although a number of recent genomic and transcriptomic analyses have detected numerous histone genes in dinoflagellates, little is known about their expression. Here in, we aimed to investigate the expression pattern of histone genes under nutritional stress, and an attempt was made to detect histone expression at the protein level in Alexandrium pacificum. The presence of histones at the mRNA level was confirmed in this study by the amplification, cloning, and sequencing of 10 different genes. Relative expression profiling of these genes under different growth conditions was determined with real-time PCR and revealed considerable levels of histone transcription in nutritionally stressed cells. We were unable to detect the expression of histones at the protein level even after immunodetection and analysis using mass spectrometry, although a histone-like protein was detected as a major nuclear component. A. pacificum expresses multiple variants of histone, and protein sequences revealed both conservation and divergence with respect to other eukaryotes. We concluded that A. pacificum maintained an active transcription of histone genes within the cell, and enhanced expression of histone genes in nutritional stress strongly suggest that histones have functional significance in dinoflagellates, although expression at the protein level was below our current detection limits, which suggests a limited role of histones in DNA packaging. Finally, the plausible regulation of histone expression at the gene and protein levels in A. pacificum is discussed.

• Journal of Photoscience
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• 제3권1호
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• pp.23-28
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• 1996

To investigate the structure and function of photosystem I, cartridge mutagenesis technique was used to inactivate the psaB gene of photosystem I. From the screen, many strains which have potential defects in photosystem I were generated. Biochemical analysis revealed that B2, one of the mutant, had a reduced amount of chlorophyll. Electron transfer activitx from photosystem II to photosystem I as oxygen uptake was the rate of 64 % of wild type. Also B2 showed a decreased photosystem I activity when measured by 77 K fluorescence emission spectrum. Particularly, immunodetection analysis showed that the B2 had reduced amount of PsaA/PsaB, but a normal range of PsaC and PsaD. Here we present a photoheterotrophic mutant for psaB gene as a unique model strain for future study of structural/functional relationship and biogenesis of photosystem I.

• Journal of Microbiology and Biotechnology
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• 제22권5호
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• pp.654-658
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• 2012

Osteoarthritis (OA) is the most common rheumatic pathology. One of the major objectives of OA research is the development of early diagnostic strategies such as those using proteomic technology. Synovial fluid (SF) in OA patients is a potential source of biomarkers for OA. The efficient and reliable preparation of SF proteomes is a critical step towards biomarker discovery. In this study, we have optimized a pretreatment method for twodimensional gel electrophoresis (2DE) separation of the SF proteome, by enriching low-abundance proteins and simultaneously removing hyaluronic acid, albumin, and IgG. SF samples pretreated using this optimized method were then evaluated by 1DE and 2DE separation followed by immunodetection of cartilage oligomeric matrix protein (COMP), a known OA biomarker, and by the identification of 3 proteins (apolipoprotein, haptoglobin precursor, and fibrinogen D fragment) that are related to joint diseases.

• BMB Reports
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• 제42권9호
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• pp.605-610
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• 2009

Plasma high-density lipoprotein cholesterol (HDL-C) levels are inversely correlated with the risk of cardiovascular disease, and are known to increase with repetitive exercise. In the current study, HDL fractions from athletes' sera were isolated and compared as a function of the type of sport (runners [n = 10], throwers [n = 10], wrestlers [n = 10], and weight lifters [n = 8]), and as an age- and gender-matched reference group (n = 14). Among athletes, HDL from runners had the strongest antioxidant activity. Immunodetection showed that runners and wrestlers had the highest levels of apoA-I and lowest levels of apoA-II in their HDL. Electron microscopy also revealed that HDL2 of runners and wrestlers were the largest in size. In conclusion, although all athlete groups had significantly better serum lipid/lipoprotein profiles than the reference group, runners and wrestlers had the most desirable lipoprotein function and structure, including antioxidant activity, HDL-associated enzyme activities and increased particle size.

• Bulletin of the Korean Chemical Society
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• 제23권10호
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• pp.1413-1431
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• 2002

For the development of immunodetection method of 4,4'-dichlorodipheny-2,2,2-trichloroethane (p,p'-DDT), a persistent and broad toxic organochlorine insecticide, various DDT derivatives were synthesized and characterized for the use of immunogens and the coating ligands for the antibody evaluation. The appropriate lengths of linkers were introduced to investigate more efficient DDT derivatives. Among these hapten derivatives, 2,2-Bis(4-chlorophenyl)acetic acid (DDA), 5,5-Bis(4-chlorophenyl)-5-hydroxypentanoic acid (DDHP) and 5,5-Bis(4-chlorophenyl)-5-chloropentanoic acid (DDCP) were conjugated with keyhole limpet hemocyanin (KLH) for the use of immunogen to produce antibodies. 6,6-Bis(4-chlorophenyl)-6-hydroxyhexanoic acid (DDHH) and 3-[6,6-Bis(4-chlorophenyl)-6-hydroxyhexanoylamino]propanoic acid (DDHHAP) in addition to above hapten derivatives were conjugated to ovualbumin (OVA) and bovine serum albumin (BSA) for the use of coating ligands to measure the titration level of antibody and the displacement of free analytes. Three matching pairs of antibodies and coating ligands were selected for the simultaneous detection of p,p'-DDT and its related compounds of DDA and 2,2-bis(4-chlorophenyl)-1,1-dichloroethylene (p,p'-DDE) by investingating the displacement of free analytes in an indirect ELISA. These were PAb #1 and coating ligand DDCP-OVA, PAb #1 and DDHHAP-OVA, and PAb #3 and DDHHAP-OVA. The most useful immunoreaction for DDT analytes were obtained using PAb #3 and coating ligand DDHHAP-OVA showing 3.4 ng/mL of lower limit of detection. These results indicated that titration level and free analytes displacement were greatly influenced by hapten derivatized and carrier proteins conjugated.

• Bulletin of the Korean Chemical Society
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• 제24권11호
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• pp.1605-1608
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• 2003

To development an immunodetection method for DDT, 1,1,1-trichloro-2,2-bis(4-chorophenyl)ethane (p,p'-DDT) and its metabolites (p,p'-DDA, p,p'-DDE, p,p'-DDD), five derivatives of DDT haptens have been synthesised and characterized as coating ligands for antibody evaluation. The appropriate lengths of linkers were introduced to investigate a matching pair of coating ligand and antibody. Among these hapten derivatives, 2,2-bis(4-chlorophenyl)acetic acid (DDA), 5,5-bis(4-chlorophenyl)-5-hydroxypentanoic acid (DDHP) and 5,5-bis(4-chlorophenyl)-5-chloropentanoic acid (DDCP) were conjugated with keyhole limpet hemocyanin (KLH) for its use as an immunogen. The bovine serum albumin (BSA) conjugates of these derivatives were prepared as a coating ligand for monoclonal antibody screening. Fifteen monoclonal antibody clones were screened using these probes. 6,6-Bis(4-chlorophenyl)-6-hydroxyhexanoic acid (DDHH) and 3-[6,6-Bis(4-chlorophenyl)-6-hydroxyhexanoylamino]propanoic acid (DDHHAP), in addition to the above hapten derivatives, were conjugated to ovualbumin (OVA) and bovine serum albumin (BSA) for their use as coating ligands to measure the titration level of the antibody and the displacement of free analytes. The indirect competitive ELISA results indicate that the titration level and free analyte displacement were greatly influenced by the DDT derivatives and carrier proteins used. Three matching pairs of monoclonal antibodies and coating ligands were selected for the DDT immunoassay: antibody clone 1A3 and coating ligand DDA-OVA, 1A1 and DDHHAP-BSA, and 1A4 and DDHP-OVA.

• 생명과학회지
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• 제9권3호
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• pp.253-261
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• 1999

Leptin gene, an obesity gene, has been known to involve in the regulation of food intake and body weight. It is also thought to be related to the glucose metabolism, insulin secretion and type II diabetes mellitus. Recently, the production of recombinant leptin protein has been attempted for the application in the treatment of obesity and the correction of hereditary obesity and type II diabetes. In the present study, leptin cDNA was cloned from mouse fat cells by RT-PCR and prokaryotic expression of leptin was attempted in order ot prepare a leptin-specific antigen. Immunization of a rabbit with the leptin-specific antigen into a rabbit resulted in the generation of leptin-specific antiserum that could be useful in the detection of leption expressed in various tissues. The sequence of leptin cDNA prepared in the present study wa identical to the previously reported one. Transformation of E. coli(DH5a) cells with the leptin cDNA-inserted translation vector, pGEX-4T-3-leptin followed by treatment with IPTG (0.1mM) resulted in the expression of a large amount of GST-leptin fusion protein with a molecular weight of 44 KDa as an inclusion body. Denaturation of the insoluble fusion protein by 8M urea, 6M guanidium-HCI or 0.1% 2-mercaptoethanol followed by a slow oxidation could not solubilize the inclusion body. The cell extract was subjected to SDS-PAGE and GST-leptin protein electroeluted from the gel was then injected into a rabbit subcutaneously for the immunization. Anti-GST-leptin rabbit antiserum which had a cross reactivity to the GST-leptin protein was generated. Leptin protein expressed in mouse brain and fat tissues was detected by Western blot immunodetection system using the antiserum generated in the present study.

• 대한치과교정학회지
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• 제32권6호
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• pp.443-453
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• 2002

본 연구의 목적은 다양한 교정용 브라켓의 표면에 형성되는 타액성 피막의 조성을 확인하고, 전타액, 악하선타액 및 이 하선타액에서 유래하는 타액성 피막의 성분을 비교하는 것이다. 네 가지 서로 다른 종류의 교정용 브라켓을 본 연구에 사용하였다. 이들은 $022{\times}028$ Roth Prescription의 상악 소구치 브라켓으로 조성은 다음과 같다: 스테인레스 스틸, 단결정 사파이어, 다결정 알루미나 및 플라스틱 브라켓. 교정용 브라켓을 각각 전타액, 이하선타액 및 악하선타액에 2시간 배양하여 타액성 피막을 형성시켰다 브라켓 피막의 타액성분은 sodium dodecyl sulfate-polyacrylamide gel electrophoresis, Western transfer method 및 면역검출법을 통해 확인하였다. 이 결과 low-molecular weight salivary mucin, ${\alpha}-amylase$, secretory IgA (sIgA), acidic proline-rich proteins, cystatins 등이 모든 브라켓의 타액성 피막에 존재하였으며, 치아우식증의 원인균인 Streptococcus mutans의 부착을 촉진시키는 타액단백질인 high-molecular weight mucin은 어떤 브라켓에도 부착하지 않았다. 그러나, 비록 동일한 타액단백질이 모든 브라켓에서 발견되었지만, 타액단백질 부착 양상은 타액의 종류 및 브라켓의 종류에 따라 양적 및 질적으로 다르게 나타났다. 특히 sIgA는 이하선타액에서 유래한 브라켓 피막에 더 많이 부착하였고, cystatins의 경우는 플라스틱 브라켓에서 유래한 브라켓 피막에 더 많이 존재하였다 본 연구는 다양한 타액단백질이 교정용 브라켓에 부착하며, 타액단백질이 타액의 출처 및 브라켓의 종류에 따라 교정용 브라켓의 표면에 선택적으로 부착함을 나타내었다.