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Preparation and Characterization of Anti-GP73 Monoclonal Antibodies and Development of Double-antibody Sandwich ELISA

  • Li, Qi-Wen;Chen, Hong-Bing;Li, Zhi-Yang;Shen, Peng;Qu, Li-Li;Gong, Lai-Ling;Xu, Hong-Pan;Pang, Lu;Si, Jin
    • Asian Pacific Journal of Cancer Prevention
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    • v.16 no.5
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    • pp.2043-2049
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    • 2015
  • Background: Serum Golgi protein 73 (GP73) as a novel and potential marker for diagnosing hepatocellular carcinoma (HCC) have been found to be elevated in HCC patients and associated with clinical variables representing tumor growth and invasiveness. The aim of this study was to prepare a pair of monoclonal antibodys (mAbs) against GP73 and develop a newly designed double-antibody sandwich enzyme-linked immunosorbent assay (s-ELISA), which would be used in the detection of serum GP73 (sGP73) as well as in the diagnosis of HCC. Materials and Methods: Produced by prokaryotic expression, the purified recombinant GP73 (rGP73), produced by prokaryotic expression, was used to immunize the Balb/c mice. Two hybridoma cell lines against GP73 were obtained by fusing mouse Sp2/0 myeloma cells with spleen cells from the immunized mice. The titers of anti-GP73 mAb reached 1:243,000. Western blotting analysis and Immunohistochemistry staining revealed that anti-GP73 mAb could recognize GP73 protein. The double-antibody s-ELISA was successfully established and validated by 119 HCC and 103 normal serum samples. Results: showed that the detection limit of this method could reach 1.56 ng/ml, and sGP73 levels in HCC group (mean=190.6 ng/ml) were much higher than those of in healthy controls (mean=70.92 ng/ml). Conclusions: Results of our study not only showed that sGP73 levels of HCC patients were significantly higher than those of healthy controls, but also indicated that the laboratory homemade anti-GP73 mAbs could be the optimal tool used in evaluating sGP73 levels, which would provide a solid foundation for subsequent clinical applications.

Production and Characterization of α-Galactosidases from Two Bacillus licheniformis Isolates (Bacillus licheniformis 분리균 2종의 α-Galactosidase 생산성과 효소특성)

  • Jin, Hyun Kyung;Yoon, Ki-Hong
    • Microbiology and Biotechnology Letters
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    • v.43 no.3
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    • pp.195-203
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    • 2015
  • Two bacterial strains, Bacillus licheniformis YB-1413 and YB-1414, producing extracellular α-galactosidase, were obtained from homemade Doenjang. On the basis of their biochemical properties, 16S rRNA sequences and random amplified polymorphic DNA patterns by polymerase chain reaction, they were found to be somewhat different from one another. α-Galactosidase productivities of the two isolates were increased by wheat bran, but drastically decreased by melibiose, raffinose and sucrose which were used as carbon sources. The enzyme productivities were increased by yeast extract as a nitrogen source with maximum levels of 1.87 U/ml for YB-1413 and 1.69 U/ml for YB-1414, respectively. The enzymes of both isolates exhibited maximum activity for hydrolysis of para-nitrophenyl-α-D-galactopyranoside (pNP-αGal) under reaction conditions of pH 6.0 and 45℃. Their hydrolyzing activities for pNP-αGal were drastically decreased by the addition of low concentrations of ribose and galactose. They were capable of hydrolyzing completely α-1,6 linked galactosyl residue in melibiose, raffinose and stachyose, which are known to be anti-nutritional factors in products of soybean and legume. In relation to the latter, the isolates YB-1413 and YB-1414 have potential applicability in improving soybean-fermented foods and the nutritional value of soybean feed.

Effect of dietary mannanoligosaccharide supplementation on nutrient digestibility, hindgut fermentation, immune response and antioxidant indices in dogs

  • Pawar, Mahesh M.;Pattanaik, Ashok K.;Sinha, Dharmendra K.;Goswami, Tapas K.;Sharma, Kusumakar
    • Journal of Animal Science and Technology
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    • v.59 no.5
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    • pp.11.1-11.7
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    • 2017
  • Background: Use of prebiotics in companion animal nutrition is often considered advantageous over probiotics because of the ease of handling, ability to withstand processing and storage etc. While most of the studies on prebiotic use in dogs have been done with processed food as basal diet, the response in relation to homemade diet feeding is not very well explored. Methods: The study was conducted to evaluate the effects of dietary mannanoligosaccharide (MOS) supplementation on nutrient digestibility, hindgut fermentation, immune response and antioxidant indices in dogs. Ten Spitz pups were divided into two groups: control (CON) with no supplementation, and experimental (MOS) wherein the basal diet was supplemented with MOS at 15 g/kg diet. All dogs were fed on a home-prepared diet for a period of 150 days. The study protocol included a digestion trial, periodic blood collection and analysis for lipid profile and erythrocytic antioxidants. Immune response of the animals was assessed towards the end of the feeding period. Results: Results revealed no significant (P > 0.05) variations in palatability score, intake and apparent digestibility of nutrients between the groups. Faecal score, faeces voided, faecal pH, concentrations of ammonia, lactate and short-chain fatty acids were comparable (P > 0.05) between the two groups. Cell-mediated immune response, assessed as delayed-type of hypersensitivity response, was significantly higher (P < 0.05) in the MOS group. The percent of lymphocyte sub-populations CD4+ and ratio of CD4+:CD8+ were also significantly (P < 0.05) higher in MOS group. The serum IgG levels were similar (P > 0.05) in both the groups. Supplementation of MOS lowered (P < 0.05) serum total- and LDL- cholesterol levels, when compared with the control group. The erythrocytic antioxidant indices were similar (P > 0.05) between the two groups. Conclusions: The results indicated that supplementation of MOS at the rate of 15 g/kg in the diet of dog augmented the cell-mediated immune response and serum lipid profile without any influences on digestibility of nutrients, hindgut fermentation and antioxidants indices.

Evaluation of Corrected Dose with Inhomogeneous Tissue by using CT Image (CT 영상을 이용한 불균질 조직의 선량보정 평가)

  • Kim, Gha-Jung
    • The Journal of Korean Society for Radiation Therapy
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    • v.18 no.2
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    • pp.75-80
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    • 2006
  • Purpose: In radiation therapy, precise calculation of dose toward malignant tumors or normal tissue would be a critical factor in determining whether the treatment would be successful. The Radiation Treatment Planning (RTP) system is one of most effective methods to make it effective to the correction of dose due to CT number through converting linear attenuation coefficient to density of the inhomogeneous tissue by means of CT based reconstruction. Materials and Methods: In this study, we carried out the measurement of CT number and calculation of mass density by using RTP system and the homemade inhomogeneous tissue Phantom and the values were obtained with reference to water. Moreover, we intended to investigate the effectiveness and accuracy for the correction of inhomogeneous tissue by the CT number through comparing the measured dose (nC) and calculated dose (Percentage Depth Dose, PDD) used CT image during radiation exposure with RTP. Results: The difference in mass density between the calculated tissue equivalent material and the true value was ranged from $0.005g/cm^3\;to\;0.069g/cm^3$. A relative error between PDD of RTP and calculated dose obtained by radiation therapy of machine ranged from -2.8 to +1.06%(effective range within 3%). Conclusion: In conclusion, we confirmed the effectiveness of correction for the inhomogeneous tissues through CT images. These results would be one of good information on the basic outline of Quality Assurance (QA) in RTP system.

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Present status of Kochujang, a fermented hot pepper soybean paste, and its globalization (고추장 산업의 현황과 세계화 제안)

  • Shin, Dong-Hwa
    • Proceedings of the Korean Society of Postharvest Science and Technology of Agricultural Products Conference
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    • 2005.09a
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    • pp.1-23
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    • 2005
  • Kochujang, a fermented hot pepper soybean paste, is a spicy condiment mixed with red pepper, boiled rice and mold fermented soybean (as koji) which is then fermented for several months. It has unique taste of hot, sweet, salty and savory flavor by all mixed ingredients by fermentation, and is one of most famous Korean traditional foods. The scale of the kochujang industry is about 200 billion won in Korea, and 7.8 million dollars and 4,500 tons in exports. Although it is still not a large part of the food industry in Korea, the scale of soy products will increase because it is a major subsidiary food in our dietary life and has grown from a homemade to a mass produced product. Kochujang has the largest market in the scale of sales among fermented soybean products and has merit because of its domestic base and its ability to grow to an international market due to its unique fermented taste. In order to make kochujang an international food, it Is necessary to study the scientific development of kochujang and the diversity of products using kochujang that are fitted to the taste of people around the world. In addition, participation in various exhibitions and food fairs supported by the government should help to promote kochujang throughout the world. In addition, these should be actively supported by the related businesses. Red pepper, a major ingredient in kochujang, has been widely studied and shown to have a positive effect on the reduction of fat, on the strengthening of the immune system, and on active biological functions. A fermented spice like kochujang has been shown to be beneficial for the body due to the process of fermentation rather than from the red pepper itself. If we prove the various biological functions of kochujang with a particular taste and flavor, it will energize a new market. It is necessary for business to make a continuous effort supported fly the scientific world and by government policy, in order to introduce this traditional food to the world. In the present day, the kochujang market is now on a firm basis and has been activated in the field of its study. Now, it is possible to achieve the birth of a secondary international market by concentrating on these efforts.

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Measurement of Tensile Properties for Thin Aluminium Film by Using White Light Interferometer (백색광간섭계를 이용한 알루미늄 박막의 인장 물성 측정)

  • Kim, Sang-Kyo;Oh, Chung-Seog;Lee, Hak-Joo
    • Journal of the Korean Society for Nondestructive Testing
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    • v.30 no.5
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    • pp.471-478
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    • 2010
  • Thin films play an important role in many technological applications including microelectronic devices, magnetic storage media, MEMS and surface coatings. It is well known that a thin film's material properties can be very different from the corresponding bulk properties and thus there has been a strong need for the development of a reliable test method to measure the mechanical properties of a thin film. We have developed an alternative and convenient test method to overcome the limitations of previous membrane deflection experiment and uniaxial tensile test by adopting a white light interferometer having sub-nanometer out-of-plane displacement resolution. The freestanding aluminium specimens are tested to verity the effectiveness of the test method developed and get the tensile properties. The specimens are 0.5 rum wide, $1{\mu}m$ thick and fabricated through MEMS processes including sputtering. 1 to 5 specimens are fabricated on Si dies. The membrane deflection experiments are carried out by using a homemade tester consisted of a motor-driven loading tip, a load cell, and 6 DOF alignment stages. The test system is compact enough to set it up beneath a commercial white light interferometric microscope. The white light fringes are utilized to align a specimen with the tester. The Young's modulus and yield point stress of the aluminium film are 62 GPa and 247 MPa, respectively.

Evaluation of the Positional Accuracy of the Delivered Beams from the Target: A Phantom Study (방사선 치료에서 치료 표적과 조사 빔의 일치 정도 평가: 팬텀 연구)

  • Kang, Sei-Kwon;Cho, Byung-Chul;Cheong, Kwang-Ho;Ju, Ra-Hyeong;Kim, Su-Ssan;Kim, Kyoung-Ju;Choi, Sang-Gyu;Bae, Hoon-Sik;Lee, Re-Na;Oh, Do-Hoon
    • Progress in Medical Physics
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    • v.17 no.4
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    • pp.192-200
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    • 2006
  • We evaluated the positional accuracy of the delivered beams to the target in a phantom by simulating the whole process of the radiation treatments Including CT scanning, planning and beam exposures with MLCs. For this purpose, a phantom was made to calibrate the alignment between the CT and the attached laser system. A new, convenient method was also devised to align the setup lasers in the treatment room. Film was used for the Identification of the delivered beam and analyzed with a homemade computer program. The positional differences between the target and the beam centers varied with the couch rotations. The accelerator we used showed a maximum discrepancy of 2.0 mm at the table angle of $295^{\circ}$. The same measurements based on the new isocenter from the Winston-Lutz test resulted in the maximum of 1.35 mm for all rotation angles. The evaluation of the differences between the target and the beam centers is useful for the treatment planning.

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A Study on Policies to Privatize Basic Environmental Facilities in Korea (우리나라 환경기초시설의 민영화 방안 모색에 관한 연구)

  • Lee, Jae-Woong;Kim, Young-Gook;Kim, Eun-Kyu;Park, Chan-Hyuk;Choi, Deuk-Su;Chung, Jae-Chun
    • Journal of the Korea Organic Resources Recycling Association
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    • v.8 no.4
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    • pp.67-77
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    • 2000
  • For successful privatization of environmental facility, a sound environmental policy is required to satisfy social motivation and economic. The primary purpose of environmental facility is not profit, but to supply a good quality service, an efficient management tool is needed to run the facility. In foreign countries, the privatization policy was originally based on the principle of complete competition achieving economic effiency. There were efficient cooperation and division of labor between civillian and public sector. First of all, the local government had independant budget. But, we found that the environmental facility have two important problems, the first is low effiency of operation and the second is comsumed to government finances as a homemade rate 50% in the technic application For a successful privatization of the environmental facility, The law of private capital introduction should be amended to promote fund raising. There are some other strategies ; overcoming of Nimbyism, research development of the privatization, field trip to the successful foreign environmental facilities. To support an efficient privatization policy, the environmental budget should be managed by cost-benefit analysis.

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Production and Characterization of Thermostable Protease from Bacillus licheniformis Isolated from Korean Traditional Soybean Paste (재래식 된장에서 분리된 Bacillus licheniformis의 내열성 Protease 특성과 생산성)

  • Bae, Young Eun;Yoon, Ki-Hong
    • Korean Journal of Microbiology
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    • v.48 no.4
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    • pp.298-304
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    • 2012
  • Among 63 Bacillus strains grown at $60^{\circ}C$ from sixteen samples of homemade Korean soybean paste, one strain was selected for producing the thermostable protease. The isolate has been identified as Bacillus licheniformis on the basis of its 16S rDNA sequence, morphology and biochemical properties. Culture filtrate of the isolate showed maximal protease activity at the reaction condition of $60-65^{\circ}C$ and pH 11. The culture filtrate retained more than 87% of initial protease activity after incubation for 30 min at $60^{\circ}C$ without substrate. In order to develop the medium composition, effects of ingredients including nitrogen sources, carbon sources, metal ions and phosphate were examined for protease production of the isolate. Lactose and soytone peptone were the most effective carbon and nitrogen source for the enzyme production. After the late logarithmic growth phase the isolate began to produce the protease, and the maximum protease productivity was reached to 550 unit/ml in the optimized medium consisting of lactose (3%), soytone peptone (1.5%), $MgSO_4$ (0.1%), $K_2HPO_4$ (0.03%), and $KH_2PO_4$ (0.03%) at 28 h of incubation.

Efficient Gene Introduction into Rice Callus by Using Particle Inflow Gun System (Particle Inflow Gun을 이용한 벼 캘러스 내의 효율적 유전자 도입)

  • Song, In-Ja;Bae, Chang-Hyu;Choi, Dae-Ock;Ryo Akashi;Lee, Hyo-Yeon
    • Journal of Plant Biotechnology
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    • v.29 no.4
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    • pp.223-228
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    • 2002
  • We have developed a homemade particle inflow gun (PIG) system which has simple operation method, low price and high gene introduction efficiency into rice callus. Rice callus were inflowed with gold particles containing DNA of a plasmid, pIG121Hm, harboring intron GUS ($\beta$-glucuronidase) gene, NPTII gene and HPT gene. For optimal GUS transient expression, the effects of parameters on DNA delivery efficiency of the PIG system was investigated by scoring transient GUS expression. The highest number of blue spots was observed at 16 mM of spemidine and 1.5 M of calcium chloride, respectively. And the amount of gold particles required for the best GUD expression was 2 mg. Optimum GUS transient expression was observed at target distance of 12 cm and helium pressure of 3.5 bar (50 psi). Gene introduction efficiency of the PIG system was observed almost similar to that of the Biolistic Gun (Bio-Rad Company). Since PIG system is simple to operate and one doesn't need disposable accessaries, the PIG system can be easily applied to various replication experiments.