• Title/Summary/Keyword: ELISA system

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Anti-adherence of Antibacterial Peptides and Oligosaccharides and Promotion of Growth and Disease Resistance in Tilapia

  • Peng, K.S.;She, R.P.;Yang, Y.R.;Zhou, X.M.;Liu, W.;Wu, J.;Bao, H.H.;Liu, T.L.
    • Asian-Australasian Journal of Animal Sciences
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    • v.20 no.4
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    • pp.569-576
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    • 2007
  • Four hundred and fifty tilapias ($6.77{\pm}0.23$ g) were assigned randomly to six groups to evaluate the feasibility of the tested antibacterial peptides (ABPs) and oligosaccharides as substitutes for antibiotics. The control group was fed with a commercial tilapia diet; other five groups were fed with the same commercial diet supplemented with konjac glucomannan (KGLM), cluster bean galactomannan (CBGAM), and three animal intestinal ABPs derived from chicken, pig and rabbit at 100 mg/kg respectively. After 21 days of feeding, growth, disease resistance, and in vivo anti-adherence were determined. Furthermore, the inhibitory effect of tested agents on adhesion of Aeromonas veronii biovar sobria (A.vbs) strain BJCP-5 to tilapia enteric epithelia in vitro was assessed by cell-ELISA system. As a result, the tested agents supplemented at 100 mg/kg show significant benefit to tilapia growth and disease resistance (p<0.05), and the benefit may be correlated with their interfering in the contact of bacteria with host mucosal surface. Although none of the tested agents did inhibit the growth of BJCP-5 in tryptic soy broth at $100{\mu}g/ml$, all of them did inhibit the adhesion of A.vbs to tilapia enteric epithelia in vivo and in vitro. In vitro mimic assays show that three ABPs at low concentrations of $25{\mu}g/ml$ and $2.5{\mu}g/ml$ have the reciprocal dose-dependent anti-adherence effect. The inhibition of ABPs may be correlated with a cation bridging and/or receptor-ligand binding, but not with hydrophobicity. The KGLM and CBGAM inhibited the adherence of BJCP-5 to tilapia enteric epithelia with dose-dependent manner in vitro, and this may be through altering bacterial hydrophobicity and interfering with receptor-ligand binding. Our results indicate that the anti-adherence of the tested ABPs and oligosaccharides may be one of the mechanisms in promoting tilapia growth and resistance to A.vbs.

Synthesis of I-125 Labelled Compound of Taxol Analogue for Radioimmunoassay (Taxol의 방사면역측정을 위한 I-125 표지화합물 합성)

  • 오옥두;금준섭;이양호;박용석;편웅범;최창운
    • Biomedical Science Letters
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    • v.3 no.2
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    • pp.125-130
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    • 1997
  • Taxol, an anticancer drug that has diterpenoid conformation, has been used as an effective chemotherapeutical agent in the treatment of breast and ovarian cancers. Because of its toxicity like other anticancer drugs, monitoring the taxol level in serum is important procedure during cancer therapy. The various monitoring methods using HPLC, ELISA, and RIA have been adopted, and RIA technique is known to be superior than other methods in trems of sensitivity and convenience. In this study, in order to develope taxol RIA system using $^{125}$I labelled antigen, first of all we synthesized taxol derivatives. 2'-hemisuccinyltaxol was obtained with about 80% yield by esterification of taxol at C-2' hydroxyl group on C-13 carbon with succinic anhydride. [$^{125}$I]iodotyramine was prepared with 58% labelling yield by radioiodination of tyramine and purified by gel chromatography. 2'-[$^{125}$I]iodotyramine-hemisuninyltaxol, $^{125}$I labelled antigen for taxol RIA, was synthesized with 96% yield from conjugation of 2'-hemisuccinyltaxol and [$^{125}$I]iodotyramine. Anti-taxol serum was produced from the rabbit immunized with 2'-hemisuccinyltaxol-BSA synthesized by 2'-hemisuccinyltaxol and BSA. The antiserum titer was determined by RIA using 2'-[$^{125}$I]iodotyramine-hemisuccinyltaxol. The titer of 1:20 was obtained with about 40% of B/T. The results suggest that taxol RIA using $^{125}$I labelled antigen can be applied to monitor the taxol level in serum.

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Determination of Appropriate Sampling Time for Job Stress Assessment: the Salivary Chromogranin A and Cortisol in Adult Females

  • Hong, Ran-Hi;Yang, Yun-Jung;Kim, Sang-Yon;Lee, Won-Young;Hong, Yeon-Pyo
    • Journal of Preventive Medicine and Public Health
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    • v.42 no.4
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    • pp.231-236
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    • 2009
  • Objectives : This study was conducted to determine the appropriate sampling time of the salivary stress markers, chromogranin A (CgA) and cortisol as objective indices of job stress assessment in adult females. Methods : The subjects were 20${\sim}$39-year-old women (13 office workers, 11 sales-service workers, and 11 college students) who were eligible for the study and free of acute and chronic medical conditions. Salivary CgA and cortisol levels were determined by enzyme-linked immunosorbent assay (ELISA). Saliva samples were collected (2 $m{\ell}$ each) at 7:00, 8:00, 10:30, 12:00, 17:30, and 22:30 on a typical day. Salivary CgA and cortisol levels, according to sampling time, were compared among the three groups using general linear model. The full version of the Korean Occupational Stress Scale (KOSS), which includes socioeconomic characteristics, health behavior, workrelated characteristics, and BMI, was used to access the subjects' job stress. Multiple regression analysis of the job stressors identified by the KOSS was performed on salivary CgA and cortisol levels. Results : The salivary CgA level peaked at 7:00 (time of awakening), then decreased and were maintained at a low level throughout the day, and increased slightly at 17:30. The salivary cortisol level increased steeply within the 1st hour after awakening, followed by a gradual decrease by 12:00, and was then maintained at a low level throughout the day. The salivary cortisol levels of subjects who worked ${\leq}$5 days per week and graduated from the university were significantly lower at 8:00 (p=0.006). The salivary cortisol levels of non-smokers were significantly lower at 7:00 p=0.040) and 8:00 (p=0.003) compared to smokers. There were no significant differences in salivary CgA and cortisol levels at 10:30 and 12:00 in general characteristics. The regression coefficients on salivary CgA level were significant with interpersonal conflict at 17:30 and job insecurity at 22:30. Regression coefficients on salivary cortisol level were significant with organizational system and total job stressors at 17:30. Conclusions : We suggest that the appropriate sampling times for the salivary stress markers, CgA and cortisol, are at 7:00 (time of awakening), 8:00 (1 hour after awakening), 17:30 (early evening), and 22:30 (before sleep).

Controlled release of nerve growth factor from heparin-conjugated fibrin gel within the nerve growth factor-delivering implant

  • Lee, Jin-Yong;Kim, Soung-Min;Kim, Myung-Jin;Lee, Jong-Ho
    • Journal of the Korean Association of Oral and Maxillofacial Surgeons
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    • v.40 no.1
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    • pp.3-10
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    • 2014
  • Objectives: Although nerve growth factor (NGF) could promote the functional regeneration of an injured peripheral nerve, it is very difficult for NGF to sustain the therapeutic dose in the defect due to its short half-life. In this study, we loaded the NGF-bound heparin-conjugated fibrin (HCF) gel in the NGF-delivering implants and analyzed the time-dependent release of NGF and its bioactivity to evaluate the clinical effectiveness. Materials and Methods: NGF solution was made of 1.0 mg of NGF and 1.0 mL of phosphate buffered saline (PBS). Experimental group A consisted of three implants, in which $0.25{\mu}L$ of NGF solution, $0.75{\mu}L$ of HCF, $1.0{\mu}L$ of fibrinogen and $2.0{\mu}L$ of thrombin was injected via apex hole with micropipette and gelated, were put into the centrifuge tube. Three implants of experimental group B were prepared with the mixture of $0.5{\mu}L$ of NGF solution, $0.5{\mu}L$ HCF, $1.0{\mu}L$ of fibrinogen and $2.0{\mu}L$ of thrombin. These six centrifuge tubes were filled with 1.0 mL of PBS and stirred in the water-filled beaker at 50 rpm. At 1, 3, 5, 7, 10, and 14 days, 1.0 mL of solution in each tubes was collected and preserved at $-20^{\circ}C$ with adding same amount of fresh PBS. Enzyme-linked immunosorbent assay (ELISA) was done to determine in vitro release profile of NGF and its bioactivity was evaluated with neural differentiation of pheochromocytoma (PC12) cells. Results: The average concentration of released NGF in the group A and B increased for the first 5 days and then gradually decreased. Almost all of NGF was released during 10 days. Released NGF from two groups could promote neural differentiation and neurite outgrowth of PC12 cells and these bioactivity was maintained over 14 days. Conclusion: Controlled release system using NGF-HCF gel via NGF-delivering implant could be an another vehicle of delivering NGF to promote the nerve regeneration of dental implant related nerve damage.

Matrix Metalloproteinase-1 and Tissue Inhibitor of Metalloproteinase-1 levels in Exudative Pleural Effusions (삼출성 흉수에서 기질 금속단백분해효소-1과 금속단백분해효소의 조직억제제-1)

  • Cho, Jae Hwa;Nam, Jeong Hyeon;Lee, Kyum Ho;Yoon, Byeong Kab;Ryu, Jeong Sun;Kwak, Sung Min;Lee, Hong Lyeol
    • Tuberculosis and Respiratory Diseases
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    • v.59 no.5
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    • pp.517-521
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    • 2005
  • Background : The balances of the proteinases and antiproteinases system have been implicated in the pathogenesis of various exudative pleural effusions. The aim of this study was to examine the matrix metalloproteinase-1 (MMP-1) and tissue inhibitor of metalloproteinase-1 (TIMP-1) levels in exudative pleural effusions. Methods : The study included 33 tuberculous effusions, 17 malignant, and 5 transudates. The pleural levels of MMP-1 and TIMP-1 were determined using a commercially available ELISA assay. Results : The group of tuberculous effusions showed higher pleural MMP-1 levels than the malignant and transudates. The pleural TIMP-1 levels of the tuberculous and malignant effusions were higher than the transudates. Conclusion : Elevated pleural MMP-1 and TIMP-1 levels were found in tuberculous effusions.

Sterol Composition and Phytoestrogen Activity of Safflower(Carthamus tinctorius L.) Seed (홍화(Carthamus tinctorius L.)씨의 sterol 및 Phytoestrogen 분석)

  • 최영주;최상욱
    • Journal of Life Science
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    • v.13 no.4
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    • pp.529-534
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    • 2003
  • This study was done to investigated the phytosterol compositions of safflower (Carthamus tinctorius L.) seed. The phytoestrogen activity was also determined using CAT-ELISA Kit in ethanol extract of safflower seed. The phytosterol of safflower seeds was identified using gas chromatography-mass spectrometry after saponification of the oils. The phytosterol content and composition of safflower seed oils were 4% and identified stigmast-5-en-3-ol (3$\beta$, 24S)-form, ${\gamma}$-sitosterol (clionasterol) with Wiley MS spectrum library. The synergistic effect of human estrogen receptor (hER) has been investigated using a minimal chimeric promoters composed of the TATA region of the adenovirus-2 major late promoter (A22MLP) and two consensus perfectly polindromic Xenopus vitellogenin A2 gene estrogen responsive elements (XVEREl19). Transient transfection experiments in tile human breast adenocarcinoma cell line MCF-7, which is known to express the estrogen receptor endogenously, revealed that phytoestrogen from Carthamus tinctorius L. acts as estrogen. We have observed the transcriptional activities stimulated methanol and ethanol extract of safflower seed in MCF-7, were 0.43 and 0.37 respectively, compared to that by $\beta$-estradiol as 1.0. Our data showed that safflower seeds have estrogenic activity methanol and ethanol extracts and ethanol lower than that of $\beta$-estradiol. This result provides the first evidence that the beneficial effect of safflower seeds may be mediated, at least in part, by the stimulating effect of phytoestrogen ell bone-protecting.

Comparison of Blood Gas, Biochemical Factors, and Hormone Concentrations, and Muscle Stress Response Factors of Chickens in General Farms and Animal Welfare Farms (일반농장과 동물복지농장 닭의 혈액 내 gas, 생화학인자, 호르몬 농도 및 근육 내 스트레스 반응인자 비교)

  • Kim, Hee-Eun;Park, Jin-Ryong;Lee, Jeong-Eun;Kang, Da-Rae;Jang, Ae-Ra;Choe, Ho-Sung;Shim, Kwan-Seob
    • Korean Journal of Organic Agriculture
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    • v.29 no.3
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    • pp.415-427
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    • 2021
  • It is generally recognized that high stocking density can increase the risk of immune dysfunction and reduction in productivity cause of elevated stress in broiler farm. The domestic livestock industry is implementing a certification system for animal welfare and livestock farms to reduce the problem of dense breeding. This study compared broiler from the general farm and animal welfare farm to confirm the effectiveness of animal welfare farms against stress. Gas concentrations in a farm of general and animal welfare farms were analyzed, blood (n=10) of the 28-day-old broiler was collected, the partial pressure of the gas in the blood was analyzed using whole blood, and biochemical factors and hormones were analyzed using the serum. Glycogen and L-lactate are analyzed using ELISA kit on finely pulverized muscles (n=10). CO2 and NH3 levels in the farm were significantly higher in general farm. Partial carbon dioxide tension (PCO2) in blood was significantly higher and partial oxygen tension (PO2) was significantly lower in the general farm. Creatinine (CREA), glucose (GLU), aspartateaminotransferase (AST), and creatinekinase (CK) in the blood were significantly higher in the general farm, and total protein (TP) and globulin (GLOB) were significantly higher in animal welfare farm. Blood corticosterone and norepinephrine were significantly higher in general farm and epinephrine was significantly higher in animal welfare farm. These results imply that animal welfare farm provides a stress-reduced breeding environment.

Studies on the regulation of Hematopoietic enhancement of Brassica campestris var narinosa., Canavalia gladiata DC semen and their combinational prescription via Jak2/STAT5/GATA1 Pathway in Sca-1+ hematopoietic stem cells (Sca-1+골수조혈세포에서 JAK2/STAT5/GATA-1 신호전달 경로를 통한 다채, 도두 그리고 두 조합물에 의한 조혈증진 조절에 관한 연구)

  • Kim, Kunhoae;Kim, Seung-Hyung;Cho, In-Sik;Kim, Han-Young;Kim, Dong-Seon;Lee, Young-Cheol
    • The Korea Journal of Herbology
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    • v.28 no.4
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    • pp.7-16
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    • 2013
  • Objectives : Brassica campestris var narinosa (BCN), Canavalia gladiata DC semen (CGD) and their combinational prescription (BCN+CGD) have been use to demonstrate to regulate hematopoiesis. In the current study, we investigated whether Brassica campestris var narinosa, Canavalia gladiata DC semen and their combinational prescription is related to hemato-potentiating function using Sca-$1^+$ hematopoietic stem cells (Sca-$1^+HSCs$) as a testing system. Methods : Sca-$1^+HSCs$ isolated from femur in C57bl/6 mice with leukopenia and thrombocytopenia induced by cyclophosphamide (CTX). Then, Real-time PCR was performed to measure the mRNA expression, ELISA and haematopoiesis-related gene (EPO, TPO, IL-3, SCF, c-kit, GM-CSF), the phosphorylation of JAK2, GATA-1 and STAT-5a/b were observed by western blot, and the numbers of $CD117^+/Sca-1^+$ cell and the number of granulocyte erythrocyte monocyte macrophage colony-forming units (CFU-GEMM) and erythroid burst forming units (BFU-E), semisolid clonogenic assay was performed. Result : When Sca-$1^+HSCs$ were treated with Brassica campestris var narinosa, Canavalia gladiata DC semen and their combinational prescription with rIL-3/rSCF, the expression of haematopoiesis-related (EPO, TPO, IL-3, SCF, c-kit, and GM-CSF) were significantly increased at the levels of mRNA as well as production in Sca-$1^+HSCs$. Additionally, CGS enhanced phosphorylation of JAK2, GATA-1, and signal transducer and activator of transcription-5a/b (STAT-5a/b) in Sca-$1^+HSCs$. Furthermore, their combinational prescription (BCN+CGD) significantly enhanced the growth rate of granulocyte erythrocyte monocyte macrophage colony-forming units (CFU-GEMM) and erythroid burst forming units (BFU-E) in vitro. Conclusion : These result suggest that Brassica campestris var narinosa (BCN) and Canavalia gladiata DC have hematopoietic enhancement via hematopoietic cytokine-mediated JAK2/GATA-1/STAT-5a/b pathway, and their combinational prescription (BCN+CGD) has superior hematopoietic enhancement to those of individual extracts.

Anti-oxidative and Anti-inflammatory Effect of Fractionated Extracts of Smilacis Glabrae Rhizoma in Human Umbilical Vein Endothelial Cell (혈관내피세포에서 토복령(土茯苓)의 항산화 및 항염증 효과)

  • Lee, Chang-Hyun;Yi, Hyo-Seung;Kim, Jae-Eun;Heo, Sook-Kyoung;Cha, Chang-Min;Won, Chan-Wook;Park, Sun-Dong
    • The Korea Journal of Herbology
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    • v.24 no.3
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    • pp.39-50
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    • 2009
  • Objectives : Smilacis glabrae rhizoma (SG) has been traditionally used as a herbal medication of musculoskeletal disorders like arthritis, pain, convulsions, and syphilis in traditional Korean medicine. This study was investigated anti-oxidative and anti-inflammatory effect of fractionated extracts of Smilacis Glabrae Rhizoma in Human Umbilical Vein Endothelial Cell (HUVEC). Methods : SG extract prepared with methanol, and then fractionated with hexane, dichloromethane, ethylacetate, n-butanol and water. Inhibitory effect of SG onto free radical generation was determined by measuring DPPH, superoxide anions and nitric oxide scavenging activities in vitro. Cytotoxic activity of extracts on RAW 264.7 cells was measured using 5-(3-caroboxymethoxyphenyl)-2H-tetra-zolium inner salt (MTS) assay. Intracelluar oxidation was analysed by DCF-DA assay. The nitric oxide (NO) production was measured by Griess reagent system. The levels of ICAM-1 and VCAM-1 expression were confirmed by western blot. And proinflammatory cytokines were measured by ELISA kit. Results : Our results indicated that fractionated extracts, especially ethyl acetate (EA) extract, significantly inhibited free radical generation, the TNF-$\alpha$-induced intracellular oxidation. Furthermore, the EA extract protected TNF-$\alpha$-induced adhesion to THP-1, expression of adhesion molecules accompanied by an attenuation of IL-6 and IL-8 formation in HUVEC. Conclusions : These results indicate that EA extract of SG have potential as an agent of atherosclerosis and other chronic inflammatory diseases including diabetes, hypertension, and arthritis.

Circulating Cytokine Levels and Changes During the Treatment in Patients with Active Tuberculosis in Korea (결핵 환자의 치료경과 중 혈청 내 Cytokine 분비와 변화)

  • Ryu, Yon-Ju;Kim, Yun-Jung;Kwon, Jung-Mi;Na, Youn-Ju;Jung, Yu-Jin;Seoh, Ju Young;Cheon, Seon Hee
    • Tuberculosis and Respiratory Diseases
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    • v.55 no.2
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    • pp.140-153
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    • 2003
  • Background : The cell-mediated immune reaction to tuberculosis infection involves a complex network of cytokines. The extent of inflammation, tissue damage and severity of the disease suggested to be determined by the balance between extent and duration of the proinflammatory cytokine response versus those of the suppressive cytokines. The systemic cytokine response in pathogenesis of tuberculosis can be assessed by measuring serum cytokine levels. Method : Serum interleukin-1 beta(IL-$1{\beta}$), IL-2, IL-4, IL-6, IL-10, IL-12(p40), tumor necrosis factor-alpha(TNF-${\alpha}$), interferon-gamma(IFN-${\gamma}$) and transforming growth factor-beta(TGF-${\beta}$) levels were measured in 83 patients with pulmonary tuberculosis, 10 patients with endobronchial tuberculosis before treatment and 20 healthy subjects by using a sandwich ELISA. In patients with pulmonary tuberculosis, they were divided into mild, moderate and far advanced group according to the severity by ATS guidelines. To compare with those of pretreatment levels, we measured serum IL-$1{\beta}$, IL-2, IL-4, IL-6, IL-10, IL-12(p40), TNF-${\alpha}$, IFN-${\gamma}$ and TGF-${\beta}$ levels in 45 of 83 patients with pulmonary tuberculosis after 2 and 6 months of treatment. Results : 1) In sera of patients with active pulmonary tuberculosis(n=83), IL-$1{\beta}$, IL-6(p<0.05), TNF-${\alpha}$, and IFN-${\gamma}$ were elevated and TGF-${\beta}$ was decreased comparing to control. IL-2, Il-12(p40), IL-4 and IL-10 were similar between the patients with tuberculosis and control. 2) In endobronchial tuberculosis, IL-6 and TNF-${\alpha}$ were elevated and TGF-${\beta}$ was decreased comparing to control. IL-12(p40) seemed to be elevated comparing to pulmonary tuberculosis. 3) Far advanced tuberculosis showed markedly elevated IL-6 and IFN-${\gamma}$ level(p<0.05). 4) The significant correlations were noted between IL-1, IL-6 AND TNF-${\alpha}$ and between IL-12, Il-2 and IL-4(p<0.01). 5) After 2 and 6 months of standard treatment, the level of IL-6 and IFN-${\gamma}$ was significantly decreased(p<0.05). Conclusion : These results showed that an altered balance between cytokines is likely to be involved in the extent of inflammation, tissue damage and severity of the disease tuberculosis. But, it should be considered diversities of cytokine response according to type of tuberculosis and immunity in clinical application and interpreting future studies.