• Title/Summary/Keyword: ELISA method

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Evaluation of the Viability of Rat Periodontal Ligament Cells after Storing at 0℃/2 MPa Condition up to One Week: In Vivo MTT Method

  • Jang, Sun Mi;Cho, Sin-Yeon;Kim, Eui-Seong;Jung, Il-Young;Lee, Seung Jong
    • Journal of Korean Dental Science
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    • v.9 no.1
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    • pp.1-8
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    • 2016
  • Purpose: The aim of this study was to evaluate the rat periodontal ligament cell viability under $0^{\circ}C/2$ MPa condition up to one week using in vivo 3-(4,5-dimethylthiazol-2-y1)-2,5-diphenyltetrazolium bromide (MTT) assay. Materials and Methods: As soon as 110 upper molar teeth of rats were extracted, they were stored in Hartman's solution under $0^{\circ}C/2$ MPa condition for 1, 2, 3, 4, and 7 days each. All specimens were treated with in vivo MTT assay and the value of optical density was measured by ELISA reader. These values were statistically analyzed by one-way ANOVA. Result: There was no statistical difference on MTT value between immediate and 1 day storage group. There were statistically significant differences between 1 day and 2 days tsorage, 2 and 3 days storage groups, respectively. Teeth of 34, and 7 days storage groups showed significantly lower MTT valuesc ompared with shorter period storage groups. Conclusion: When the MTT values were substituted in standard curve, 1 day storage group at $0^{\circ}C/2$ MPa condition showed 68% cell viability when compared with immediate group. It dropped to 13% at 2 days, and to less than 5% at 3 days or more.

Tumor Surpressor Gene Therany, and Natural Product with Vectors[Aoenouirus, Aoenn associated virus] in Human Papilloma virus (HPV[Human papilloma virus]유래 바이러스 벡터[Adenovirus, Adeno associated virus]를 이용한 암 억제유전자치료법과 자연산물에서의 암 억제 효과)

  • 천병수;노민석;유종수;김준명
    • KSBB Journal
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    • v.16 no.6
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    • pp.579-591
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    • 2001
  • The cell growth inhibitor effect of cervical cancer cells was investigated by liposome mediated transfection (pRcCMVp53/lipofectin) and by transfection using adenovirus (AdCMVp57). The papilloma virus cancer cell lines we used in this study were HPV16 positive, having inhibiter gene, wild p53 gene, CaSki, SiHa, HPV18 positive HeLa, HeLaS3 and HPV negative C33A, HT3. LacZ gene of E.coli was used as the marker gene for the transfection efficiency. The effect on the inhibition of tumor cell growth was measured by cell count and cell viability though ELISA analysis and MTT assay. The inhibition of tumor cell growth was confirmed by measuring each assay for six days, comparing with the normal control cell growth. The cell growth of cervical cancer calls by transfection was significantly reduced and showed tittle differences among the cell lines. To eliminate the potential problem of Ad(adenovirus) contamination during rAAV production, rAAV can be produced by a triple transfection of vector plasmic, packaging plasmid, and adenovirus helper plasmid. To examine the helper functions of Ad plasmids on the production of rAAV vector, we carried out cotransfection of three plasmids, AAV vector, packaging construct, and Ad helper plasmids. The optimized transfection condition for calcium phosphate method is 25ug of total DNA per 10-cm-diameter plate of 293 cell. We found that rAAV yields peaked at 48hr after Ad infection. The titer of rAAV was measured by the dot blot analysis to measure the number of particles/ml based on the quantification of viral DNA. Recent1y, Kombucha(fungi) was identified as a very potent antileukefic agent. In the present study, effect of natural toxin(plankton) and Kombucha is PSP(GTXI-3, neoSTX), on various MTT assay cervical cancer cell line. Toxin(GTX 1-3, neoSTX) also inhibited the proliferation in primary cervical cancer calls in a dose-dependent toxin concentration. These results showed that toxin was very potent in inhibiting the proliferation of cervical cancer calls in vitro. Toxins and Kombuoha exhibited a dose dependent inhibition of cellular proliferation in cancer cell line.

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Peri-implant crevicular fluid levels of cathepsin-K, RANKL, and OPG around standard, short, and mini dental implants after prosthodontic loading

  • Alan, Raif;Marakoglu, Ismail;Haliloglu, Seyfullah
    • Journal of Periodontal and Implant Science
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    • v.45 no.5
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    • pp.169-177
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    • 2015
  • Purpose: Despite the high success rates of endosseous dental implants, their placement is restricted according to the height and volume of bone available. The use of short or mini dental implants could be one way to overcome this limitation. Thus, this study aimed to compare standard, short, and mini dental implants with regard to associated clinical parameters and peri-implant crevicular fluid (PICF) levels of cathepsin-K (CTSK), RANK ligand (RANKL), and osteoprotegerin (OPG), after prosthodontic loading. Methods: A total of 78 non-submerged implants (Euroteknika, $Aesthetica^{+2}$, Sallanches, France) were installed in 30 subjects (13 male, 17 female; range, 26-62 years) who visited the clinic of the Periodontology Department, Faculty of Dentistry, Selcuk University. Sampling and measurements were performed on the loading date (baseline) and 2, 14, and 90 days after loading. Assessment of the peri-implant status for the implant sites was performed using the pocket probing depth (PPD), modified plaque index, modified gingival index, modified sulcular bleeding index, and radiographic signs of bone loss. PICF samples collected from each implant were evaluated for CTSK, RANKL, and OPG levels using the ELISA method. Keratinized tissue and marginal bone loss (MBL) were also noted. Results: Clinical parameters statistically significantly increased in each group but did not show statistical differences between groups without PPD. Although implant groups showed a higher MBL in the upper jaw, only the standard dental group demonstrated a statistically significant difference. At 90 days, the OPG:sRANKL ratio and total amounts of CTSK for each group did not differ from baseline. Conclusions: Within the limitations of this study, both short and mini dental implants were achieving the same outcomes as the standard dental implants in the early period after loading.

Alteration of Plasma ${\beta}$-Nerve Growth Factor Concentration in Depressed Patients with Suicidal Attempt (자살을 시도한 우울증 환자에서 혈장 ${\beta}$-Nerve Growth Factor 농도의 변화)

  • Shim, Se-Hoon;Won, Seong-Doo;Lee, Bun-Hee;Han, Chang Su;Yang, Jong-Chul;Kwon, Young-Joon;Kim, Yong-Ku
    • Korean Journal of Biological Psychiatry
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    • v.13 no.2
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    • pp.95-102
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    • 2006
  • Object : Nerve growth factor(NGF) is a protein involved in neuronal survival and plasticity in the central nervous system, which might play an important role in stress, depression and suicide. This study was performed to determine whether there is an alteration in plasma NGF concentrations in depressed patients with suicidal attempt. Methods : The subjects were 32 depressed patients who attempted suicide and admitted in emergency room. Forty-four hospitalized non-suicidal depressive patients and the 30 normal controls were closely matched with the suicidal group in terms of age and sex. Individuals in all 3 groups were evaluated independently by a semi-structured interview for the purpose of establishing a DSM-IV criteria diagnosis. The severity of depressive symptoms was evaluated using Hamilton depression rating scale(HDRS). The severity of the suicidal behavior was evaluated by Weisman and Worden's risk-rescue rating(RRR) system and the Lethality Suicide Attempt Rating Scale(LSARS). Plasma NGF level was measured by the enzyme linked immunosorbent assay(ELISA) method. Results : There were no statistically significant differences of the plasma NGF levels among groups. LSARS and RRR did not reveal any significant correlation with ${\beta}$-NGF level in suicidal depressive patients. Conclusion : This study do not support an association between ${\beta}$-NGF and suicidal depression. However it is necessary to investigate this association through other route such as postmortem brain.

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The Effects of Juglandis Semen Extract on the Experimental Asthma induced by Ovalbumin (호도(胡桃) 물추출물이 Ovalbumin으로 유도된 천식동물모델에 미치는 영향)

  • Kim, Sang-Chan;Byun, Sung-Hui
    • Herbal Formula Science
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    • v.13 no.1
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    • pp.145-159
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    • 2005
  • Objective : Juglandis Semen has a function that to invigorate the lung and kidney. And It is commonly used as a supporting agent in the treatment of coughing and bronchitis. This study was performed to investigate the effect of oral administration of Juglandis Semen Extract (JSE) against the experimental asthma induced by ovalbumin. Methods : Asthma was induced to Balb/c mouse by i.p. injection and aerosol immunization with ovalbumin. It was observed the change of the cell number in the BAL fluid. Concentrations of IL-4, IL-5 in splenoc yte were assessed by ELISA, IgG and IgE from serum were calculated by same method. Results : 1. Number of macrophage in BAL fluid was significantly decreased in JSE group compared with control group, but not eosinophil and lymphocyte. 2. Levels of IgG and IgE in serum were significantly decreased in JSE group compared with control group, respectively. 3. Concentration of IL-4 in culture supernatant of splenocyte was significantly decreased in JSE group compared with control group, but there was no significant in IL-5. Conclusion : We found that the effect of JSE extract in asthma was implicated in reduction of IL4released from Th2 cell, and decreases of IgG and IgE from plasma cell. These findings suggest that JSE can produce anti-asthmatic effect, which may play a role in allergen-induced asthma therapy.

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A Rapid and Sensitive Screening System for Human Type I Collagen with the Aim of Discovering Potent Anti-Aging or Anti-Fibrotic Compounds

  • Hashem, Md. Abul;Jun, Kyu-Yeon;Lee, Eunyoung;Lim, Soyun;Park Choo, Hea-Young;Kwon, Youngjoo
    • Molecules and Cells
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    • v.26 no.6
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    • pp.625-630
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    • 2008
  • This study was undertaken with the aim of developing an easy and quick means of analyzing the effect of various compounds on the synthesis and secretion of human type I collagen at the protein level. A modification of the ELISA method was used on HFF-1 cells. For the proof of concept, we used thirteen compounds most of which are known to be antioxidants. Each compound was tested at concentrations of 0, 10 and $100{\mu}m$ on HFF-1 cells for 24 h. Thirteen sets of experiments for each compound were performed in ANOVA with three replicates. Duncan multiple range test (DMRT) was used to compare the mean values obtained from the treatment groups. From the results it was concluded that Vitamin C, undecylenic acid, conjugated linoleic acid, glycolic acid, and citric acid at $100{\mu}m$ concentration could be used for anti-wrinkling or protection from premature aging, which requires enhancement of collagen synthesis. Lactic acid, EGCG, resveratrol, and retinol that can inhibit collagen synthesis effectively in a dose-dependent manner may be used for anti-fibrosis treatment purposes.

Development of a Specific antibody for the Detection of Ice Nucleation-Active Bacteria (빙핵세균의 검출을 위한 특이적 항혈청 개발)

  • Lee, Ung;Kwon, Mi-Kyung;Seong, Ki-Young;Cho, Baik-Ho;Kim, Ki-Chung
    • Plant Disease and Agriculture
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    • v.5 no.1
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    • pp.27-33
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    • 1999
  • Frost injury of crops is closely related to the epiphytic population dynamics of ice nucleation-active (INA) bacteria, and the injury can be reduced by decreasing the INA bacterial population. In order to predict the epiphytic population of INA bacteria on crops, a rapid and accurate detection method has to be developed. In the previous report, we produced some antibodies against INA proteins purified from the outer membrane of INA bacteria. However it was difficult to produce the antibodies because the purification procedures of the INA proteins were complicated, and the final yield was too low. We designed a specific peptide from the N-terminal region of INA protein by computer analysis and synthesized the peptide in vitro in this experiment. The peptide sequence was Asp-Ser-Por-Leu-Ser-Leu-His-Ala-Asp, that is corresponding to the highly conserved region in several INA proteins, with predicted beta turn, coiling, and hydrophilic region. A polyclonal anti-INA peptide antiserum produced specifically recognized INA bacteria as few as 10 colony-forming units (CFU) in the ELISA reactions and did not respond to other non-INA bacteria. Serological specificity of the anti-INA peptide antiserum will facilitate the forecasting of the INA bacterial population dynamics on crops.

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Presumptive Diagnosis of Mycoplasma pneumoniae Pneumonia in Children (소아에서 마이코플라즈마 폐렴의 예기적 진단)

  • Lee, Chang Eon;Park, Su Jin;Kim, Won Duck
    • Journal of Yeungnam Medical Science
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    • v.29 no.2
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    • pp.89-95
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    • 2012
  • Background: As Mycoplasma pneumoniae pneumonia has increased in Korea, its relevance to infants, toddlers, and adolescents has magnified as well as. However, it is difficult to perform the serological test and PCR test routinely for diagnosis in actual clinical practice. Thus, the authors conducted this study to help clinicians do presumptive diagnosis of Mycoplasma pneumoniae pneumonia using clinical, radiological, and hematological findings. Methods: The study population consisted of 224 children between 1 month and 14 years old, hospitalized for radiographically confirmed pneumonia. Patients were divided into two groups of 100 children with Mycoplasma pneumoniae pneumonia, as diagnosed using the ELISA method. Groups with negative result in Mycoplasma IgM antibody test were classified into the viral group (98 patients with respiratory virus) and the bacterial group (46 patients with the bacteria detected in the blood sputum culture or antibiotic treatment except macrolide improved the patient's condition). These groups were compared and analyzed using clinical, hematological, and radiographic differences and scoring system. Results: Clinical, hematological, and radiographic characteristics of Mycoplasma pneumoniae pneumonia have shown the intermediate level results between bacterial pneumonia and viral pneumonia. In terms of scoring system, the mean score of Mycoplasma pneumoniae pneumonia was 4.23, which was the intermediate level between bacterial pneumonia (mean score=6.67) and viral pneumonia (mean score=1.48). Conclusion: Results suggest that the combination of the scoring system information can increase the accuracy in the diagnosis even if they may have difficulties on diagnosis, because clinical manifestations, hematological, and radiographic findings are nonspecific.

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Regulation Roles of MICA and NKG2D in Human Renal Cancer Cells

  • Jia, Hong-Ying;Liu, Jun-Li;Yuan, Ming-Zhen;Zhou, Cheng-Jun;Sun, Wen-Dong;Zhao, Jing-Jie;Wang, Jue;Liu, Ling;Luan, Yun
    • Asian Pacific Journal of Cancer Prevention
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    • v.16 no.9
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    • pp.3901-3905
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    • 2015
  • Objective: Our aim was to investigation the roles of MHC class I chain-related gene A(MICA) and natural killer cell group 2D(NKG2D) in human renal cancer cells. Materials and Methods: The expression of membrane MICA (mMICA) on renal cells and NKG2D on NK cells were detected by flow cytometry (FCM); the content of sMICA were detected by enzyme linked immunosorbent assay (ELISA) and the distribution of mMICA on renal tumor tissues by immunohistochemistry; the interaction between MICA and NKG2D was observed by antibody closed method. Results: Our results showed that the expression of mMICA in renal cancer tissues was significantly higher than in controls, where the soluble MICA was not expressed. Cytotoxic activity of NK cells was significantly reduced after exposure to NKG2D and MICA antibodies (P<0.05), and serum containing sMICA can obviously lower the function of NKG2D (P<0.05). Conclusions: The interaction of mMICA and NKG2D play important roles in mediation of cytotoxicity of NK cells in RCC. On the other hand, sMICA may mediate tumor immune escape through down- regulated NKG2D expression.

SNPs of Excision Repair Cross Complementing Group 5 and Gastric Cancer Risk in Chinese Populations

  • Yang, Wan-Guang;Zhang, Shan-Feng;Chen, Ju-Wu;Li, Li;Wang, Wan-Peng;Zhang, Xie-Fu
    • Asian Pacific Journal of Cancer Prevention
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    • v.13 no.12
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    • pp.6269-6272
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    • 2012
  • We conducted a case-control study to determine the association between several potential SNPs of excision repair cross complementing group 5 (XPG) and gastric cancer susceptibility, and roles of XPG polymorphisms in combination with H.pylori infection in determining risk of gastric cancer. In our study, we collected 337 newly diagnosed gastric cancer cases and 347 health controls. Three SNPs of XPG, rs2296147T>C, rs2094258C>T and rs873601G>A, were genotyped using the Taqman real-time PCR method with a 7900 HT sequence detector system. H. pylori infection was diagnosed by ELISA. By multivariate logistic regression analysis, the rs2296147 CC genotype was associated with a decreased risk of gastric cancer (OR=0.52, 95% CI=0.27-0.97), and rs2094258 TT was associated with elevated risk (OR=2.13, 95% CI=1.22-3.35). Positive H.pylori individuals with rs2094258 TT genotypes demonstrated increased risk of gastric cancer (OR=2.13, 95% CI=1.22-3.35), while rs2296147 CC was associated with lower risk among patients with negative H.pylori (OR=0.45, 95%CI=0.22-0.89). Our findings suggested that XPG polymorphisms might contribute to risk of gastric cancer among Chinese populations, but the effect needs to be further validated by larger sample size studies.