• Title/Summary/Keyword: sandwich-ELISA

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Liripois Tuber contributes to the chemotaxis of eosinophils and secretion of cytokines in A549 human epithelial cells (맥문동(麥門冬)이 천식유발 cytokine 분비와 호산구 chemotaxis에 미치는 영향)

  • Jung, Hae-Jun;Jung, Gee-Jae;Jung, Sung-Ki;Rhee, Hyung-Koo
    • The Journal of Korean Oriental Chronic Disease
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    • v.10 no.1
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    • pp.1-20
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    • 2005
  • Background and Objective: The importance of the presence of eosinophils in the airways of patients with fetal asthma has long been recognized, but the mechanism by which these cells are recruited and retained in the lung are only now being elucidated. Eotaxin is a potent and specific eosinophil chemoattractant that is mobilized in the respiratory epithelium after allergic stimulation. Material and Methods: We used water extracts of Liripois Tuber and pulmonary epithelial cell lines A549(human typeII-like epithelial cells) and human eosinophils. We estimated cytotoxic effects of Liripois Tuber via MTS assay, and estimated the effects of Liripois Tuber on chemokines from prestimulated A549 cells by sandwich ELISA and RT-PCR. We conducted chemotaxis assay on prestimulated eosinophils treated with Liripois Tuber. Result: In this study we demonstrated that $TNF-{\alpha}$, IL-4 and $IL-1{\beta}$ induced the accumulation of chemokines mRNA in the pulmonary epithelial cell lines A549 in dose-dependent manner. Chemokines were inhibited by Liripois Tuber in dose-dependent manner. The eosinophil migration was inhibited at high concentration of Liripois Tuber. Conculusion: These findings indicate that the supression of the expression of chemokines can be accomplished by Liripois Tuber treatment, raising the possibility that Liripois Tuber might be of therapeutic value in diseases such as asthma.

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Development of Evaluating Ways for the Efficacy of Anti-VEGF Biopharmaceuticals (VEGF 제어의약품의 효능 평가법 개발)

  • Nam, Eun-Hee;Jeon, Seong-Hyun;Lee, Wha-Jung;Seo, Dong-Wan;Kim, Pyeung-Hyeun
    • IMMUNE NETWORK
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    • v.7 no.4
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    • pp.203-208
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    • 2007
  • Background: Angiogenesis mediated by VEGF constitutes a new target for anti-cancer therapy which has explored through different ways of intervention aiming at the blocking of the tumoral angiogenesis. In the present study, we developed the assays by which efficacies of anti-VEGF inhibitor candidates are evaluated at the various levels. Methods & Results: First, we developed two sandwich ELISAs using coated anti-VEGF Ab and soluble Flt-1 receptor fusion protein (sFlt-1/Fc). As low as 200 pg/ml of hVEGF diluted in human sera was detectable by these assays. In addition, we found that VEGF inhibitors ($2{\mu}g/ml$ of either anti-VEGF Ab or sFlt-1/Fc) completely block 5 ng/ml VEGF in these ELISAs. Subsequently, two bioassays, wound healing and HUVEC tube formation assays, revealed that anti-VEGF Ab $(1{\mu}g/ml)$ & sFlt-1/Fc Ab $(1{\mu}g/ml)$, or SU5416 (VEGFR tyrosine kinase inhibitor, $1{\mu}M$) prevents the activity of VEGF $(1{\sim}10ng/ml)$. Finally, secretion of MMP-9 by VEGF-stimulated macrophages was abolished by treatment of anti-VEGF Ab $(1{\mu}g/ml)$ in gelatin zymography. Conclusion: ELISAs together with bioassays developed in this study are appropriate for evaluation of the efficacy of inhibitors of VEGF.

Insights into the Incidence of Watermelon chlorotic stunt virus Causing Yellowing Disease of Watermelon in Western and Southwestern Regions of Saudi Arabia

  • Ahmad, M.H.;Shakeel, M.T.;Al-Shahwan, I.M.;Al-Saleh, M.A.;Amer, M.A.
    • The Plant Pathology Journal
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    • v.34 no.5
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    • pp.426-434
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    • 2018
  • During the spring season of 2014, a total of 148 melon and watermelon leaf samples were collected from symptomatic and asymptomatic plants in the western and southwestern regions of Saudi Arabia and were tested for the presence of Watermelon chlorotic stunt virus (WmCSV) and other suspected cucurbit viruses by double antibody sandwich enzyme-linked immunosorbent assays. Ninety-eight samples were found to be positive for the presence of WmCSV, nine samples were positive for the presence of Cucurbit yellow stunting disorder virus (CYSDV), and 22 showed a mixed infection with both WmCSV and CYSDV. No other cucurbit viruses were detected in any of the samples. Host range experiments revealed that eight out of fourteen tested plant species were susceptible to WmCSV. PCR products of approximately 1.2 kb were obtained after amplification using primers specifically targeting the coat protein region of WmCSV. Positive PCR results were confirmed by dot blot hybridization. Coat protein gene sequences from eleven WmCSV isolates indicated that the highest identity was between the 104WMA-SA isolate from the Wadi Baish location and a previously reported isolate from the AL-Lith location in Saudi Arabia. The lowest identity was observed between the 42WMA-SA isolate and an isolate from Palestine.

Influence of Serum VEGF Levels on Therapeutic Outcome and Diagnosis/Prognostic Value in Patients with Cervical Cancer

  • Du, Ke;Gong, Hong-Ying;Gong, Zhi-Min
    • Asian Pacific Journal of Cancer Prevention
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    • v.15 no.20
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    • pp.8793-8796
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    • 2014
  • Objective: To explore the influence of serum vascular endothelial growth factor (VEGF) level on therapeutic outcome and diagnosis/prognostic value in patients with cervical cancer. Materials and Methods: A total of 37 patients diagnosed with cervical cancer by biopsy were selected and treated with concurrent chemoradiotherapy. Double-antibody sandwich enzyme-linked immunosorbent assay (ELISA) was adopted before treatment to assess VEGF levels, and its relationships with clinicopathological features and short-term therapeutic effects were analyzed. Results: The median VEGF level in 37 patients before treatment was 647.15 (393.35~1125.16) pg/mL. Serum VEGF levels in patients aged <50 years, in International Federation of Gynecology and Obstetrics (FIGO) stage IIIa~IVa, with lymph node metastasis and tumor size >4 cm were significantly increased (P<0.05). The complete remission (CR) rate was 48.7% (18/37), partial remission (PR) rate was 35.1% (13/37), stable disease (SD) rate was 13.5% (5/37) and progressive disease (PD) rate was 2.70% (1/37), so the objective remission rate (ORR) after treatment was 83.8% (31/37). Logistic regression analysis showed that tumor size and serum VEGF level before treatment were independent risk factors affecting the therapeutic outcome, and the higher the level of serum VEGF, the worse the prognosis when tumor size>4 cm. Some 56.8% of patients manifested with myelosuppression, 37.8% with leucopenia, 24.3% with thrombocytopenia, 5.41% with diarrhea, 46.0% with nausea and vomiting, 21.6% with hair loss and 8.11% with hepatic and renal injury during the treatment. Conclusions: Serum VEGF level may reflect the degree of malignancy of cervical cancer and predict therapeutic effect, which is of great importance to cancer diagnosis and prognosis.

Anti-inflammatory Effects of the Water Extract of Phyllostachys Folium via NF-κB Inhibition (죽엽 열수추출물의 염증억제 효과)

  • Son, Jin Won;Park, Sang Mi;Jung, Ji Yun;Hwangbo, Min;Cho, Il Je;Jung, Tae Young;Park, Chung A;Kim, Sang Chan;Jee, Seon Young
    • Herbal Formula Science
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    • v.24 no.4
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    • pp.259-269
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    • 2016
  • Objectives : Phyllostachys Folium is leaves of Phyllostachys nigra var. henesis $S_{TAPF}$. In the East Asian traditional medicine, the herb has been used to treat nasal bleeding, dysuria, epilepsy and etc. The present study was conducted to evaluate the anti-inflammatory effects of the Phyllostachys Folium water extracts (PFE) in vitro and in vivo model. Methods : Cell viability was measured by MTT assay after the treatment of PFE and NO production was monitored by measuring the nitrite content in culture medium. iNOS, COX-2, $I{\kappa}B$, $p-I{\kappa}B{\alpha}$ amd $NF{\kappa}B$ were detected by immunoblot analysis, and levels of cytokine were analyzed by sandwich ELISA kit. Anti-edema effect of PFE was determined in the carrageenan-induced paw edema model in rats. Results : LPS increased NO and cytokines levels compared with control, these increases were attenuated by PFE. In addition, LPS-induced pro-inflammatory proteins such as iNOS, COX-2 were down regulated by PFE. These anti-inflammatory effect of PFE results from inhibition of phosphorylation of $I{\kappa}B$ and translocation of $NF-{\kappa}B$. Conclusion : These results show that PFE has some anti-inflammatory effects which might play a role in gram-negative bacterial infection inflammation and $NF{\kappa}B$ activated diseases.

Lonicerae Flos contributes to the chemotaxis of eosinophils and secretion of cytokines in A549 human epithelial cells (금은화(金銀花)가 천식(喘息)유발 cytokine 분비와 호산구 chemotaxis에 미치는 영향)

  • Chung, Kwang-Jin;Jung, Sung-Ki;Rhee, Hyung-Koo;Jung, Hee-Jae
    • The Journal of Internal Korean Medicine
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    • v.26 no.1
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    • pp.129-142
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    • 2005
  • Objective : Asthma is known as chronic airway inflammatory disease. This inflammation is conducted by various inflammatory cells including eosihophil. Chemotaxis is one way that circulating inflammatory cells invade a specific lesion. This study examines the degree to which Lonicerae Flos inhibits eosinophil chemotaxis at pulmonary epithelium after allergic stimulation. Material and Methods : Water extracts of Lonicerae Flos and pulmonary epithelial cell lines A549(human type II-like epithelial cells) and human eosinophils were used. Cytotoxic effects of Lonicerae Flos via MTS assay were estimated, as well as the effects of Lonicerae Flos on chemokines from prestimulated A549 cells by sandwich ELISA and RT-PCR. Chemotaxis assay was conducted on prestimulated eosinophils treated with Lonicerae Flos. Result : In this study $TNF-{\alpha}$ and IL-4, $IL-l{\beta}$ were seen to induced the accumulation of chemokines mRNA in the pulmonary epithelial cell lines A549 in a dose-dependent manner. Chemokines were inhibited by Liripois Tuber in a dose-dependent manner and especially, IL-8 and ICAM-l were inhibited considerably at $100\;{\mu}g/ml$ concentration of Lonicerae Flos. The eosinophil migration is inhibited in high concentration of Lonicerae Flos in a dose-dependent manner. Conclusion : These findings indicate that the supression of the expression of chemokines can be accomplished by Lonicerae Flos treatment, raising the possibility that Lonicerae Flos might be of therapeutic value in diseases such as asthma.

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Moutan Cortex Radicis contributes to the chemotaxis of eosinophils and secretion of cytokines in A549 human epithelial cells (목단피(牧丹皮)가 천식(喘息)유발 cytokine 분비와 호산구 chemotaxis에 미치는 영향)

  • Moon, Sung-Hun;Jung, Sung-Ki;Rhee, Hyung-Koo;Jung, Hee-Jae
    • The Journal of Internal Korean Medicine
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    • v.26 no.1
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    • pp.199-212
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    • 2005
  • Objective : Airway inflammation is now regarded as a defining feature of asthma. The importance of eosinophits in the airway inflammation of asthma patients is widely recognized, and eosinophils mobilization in the respiratory epithelium is activated by chemoattractants and cytokines. This study was designed to examine the extent of the ability of Moutan Cortex Radicis to inhibit eosinophil chemotaxis of pulmonary epithelium after allergic stimulation. Material and Methods : Water extracts of Moutan Cortex Radicis and pulmonary epithelial cell lines A549(human type II-like epithelial cells) and human eosinophils were used. Cytotoxic effects of Moutan Cortex Radicis were estimated via MTS assay, and the effects of Moutan Cortex Radicis on chemokines from prestimulated A549 cells were estimated by sandwich ELISA and RT-PCR. Chemotaxis assay on prestimulated eosinophils treated with Moutan Cortex Radicis. was conducted Result : In this study we demonstrated that $TNF-{\alpha}$ and IL-4, $IL-1{\beta}$ induced the accumulation of chemokines' mRNA in the pulmonary epithelial cell lines A549 in a dose-dependent manner. Chemokines of eotaxin, ICAM-1, YCAM-1, IL-8, IL-16 were inhibited by Moutan Cortex Radicis in a dose dependent manner, but RANTES showed no inhibition due to Moutan Cortex Radicis. Eosinophil migration was inhibited at high concentrations of Moutan Cortex Radicis. Conculusion : These findings are indicative of supression of chemokines accomplished by Moutan Cortex Radicis treatment, demonstrating the potential therapeutic value of Moutan Cortex Radicis for treating diseases such as asthma.

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Effects of conjugated linoleic acid and selenium on body composition and serum cholesterol concentration in broiler chickens (CLA와 selenium의 투여가 닭의 체구성과 혈중 콜레스테롤 수준에 미치는 영향)

  • Jun, Ho-Jin;Kim, Jin-Young;Lim, Joa-Jin;Lee, Soo-Han;Lee, Byeong-Han;Jeong, Soon-Wuk;Park, Hee-Myung;Chung, Byung-Hyun
    • Korean Journal of Veterinary Research
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    • v.45 no.3
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    • pp.429-438
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    • 2005
  • The purpose of this study was to examine whether conjugated linoleic acid and selenium supplementation in broiler chicken diets would be effective, enhance indices of immune status and body weight, and modulate serum lipid concentration. Forty Hyline brown chickens, 1 weeks of age, were divided into 5 groups of 8 chickens. Chickens were fed the experimental diets supplemented with 1% CLA (conjugated linoleic acid; Group 1), 1% CLA + selenium (Group 2), 1% safflower-seed-oil as LA (Group 3), 1% safflower-seed-oil as LA + selenium (Group 4) or nothing (Control) for 4 weeks. After 4 weeks, serum, liver, spleen and abdominal fat were taken. Measurement of total immunoglobulin were executed using sandwich ELISA. Weight ratio of liver to body showed that the group fed with CLA were significantly higher than the group fed with CLA + selenium. Weight ratios of spleen and fat to body showed no significantly differences. In concentrations of serum total cholesterol and HDL-cholesterol, the group fed with CLA showed significantly higher values than that fed with CLA + selenium. In concentrations of serum triglyceride and LDL-cholesterol there were no significantly differences between the treatment groups. In conclusion, supplementation of CLA with selenium protected hepatomegaly and reduced level of serum total cholesterol and HDL-cholestererol in chickens.

Alteration of Porcine Serum Albumin Levels in Pork Meat by Marination in Kiwi or Pineapple Juice and Subsequent Pan Broiling

  • Moon, Sung-Sil;Kim, Dongwook;Kim, Il-Suk;Ham, Jun-Sang;Park, Beom-Young;Jang, Aera
    • Food Science of Animal Resources
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    • v.34 no.3
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    • pp.355-361
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    • 2014
  • This study was conducted to evaluate the changes in porcine serum albumin (PSA), a major allergen, which occur when raw pork ham is marinated with kiwi or pineapple juice, and/or when the ham is pan broiled at $300^{\circ}C$ for 4 min after marination. In this study, raw pork ham was soaked for 4 h or 8 h in marinades containing commercial marinating sauce only, commercial marinating sauce and 7% kiwi juice, or commercial marinating sauce and 7% pineapple juice. When the meat was marinated and then pan-broiled, pork ham meat protein was significantly denatured and hydrolyzed, and the level of PSA in the meat was significantly reduced. The PSA contents of pork broiled without marination, pork that had been marinated in commercial marinating sauce alone, pork that had been marinated in commercial marinating sauce with kiwi juice, and pork that had been marinated in commercial marinating sauce with pineapple juice, were 95.4, 43.3, 14.3, and 5.4 ng/mL, respectively (p<0.05). Marinating with pineapple juice was more effective than marinating with kiwi juice; and marination for 8 h was more effective than marinating for 4 h. These results indicate that the level of PSA in pork ham is effectively reduced, when the meat is first marinated in sauces that contain kiwi or pineapple extracts for 8 h, rather than 4 h, and then cooked. Further study is needed to determine whether marinated pork meat reduces allergenicity in vivo, as well.

Simultaneous Detection of Three Tobamoviruses in Cucurbits by Rapid Immunofilter Paper Assay

  • Park, Gug-Seoun;Kim, Jae-Hyun;Chung, Bong-Nam;Kim, Hyun-Ran;Park, Yong-Mun
    • The Plant Pathology Journal
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    • v.17 no.2
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    • pp.106-109
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    • 2001
  • A multi-rapid immunofilter paper assay (multi-RIPA) system was prepared for simultaneous diagnosis of three Tobamoviruses, Cucumber green mottle mosaic virus (CGMMV), Kyuri green mottle mosaic virus (KGMMV), and Zucchini green mottle mosaic virus (ZGMMV) in cucurbitaceous crops. Each of these viruses was specifically detected by the multi-RIPA from cucumber, watermelon, zucchini, and bottle gourd inoculated with the three Tobamoviruses singly or in combination. The three viruses could infect cucumber, watermelon, and bottle gourd ; however, CGMMV could not infect zucchini as the latex-coated CGMMV antibody showed a negative reaction in the multi-RIPA of the virus-infected plant extract. When the minimum detection level of multi-RIPA was compared with that of double antibody sandwich-enzyme-linked immunosorbent assay (DAS-ELISA) using CGMMV, the latter was 10 times more sensitive than the former. The detection limit of the multi-RIPA for the purified CGMMV was 50 ng/ml. In a survey of the threeviruses in cucurbits growing in commercial fields in 1999 and 2000, CGMMV was detected in watermelon and cucumber, and ZGMMV was detected only in zucchini growing in plastic houses at the suburbs of Chonju, Korea. However, KGMMV was not found in the commercially growing cucurbit crops in our study, The results suggest that the multi-RIPA can be a simple, rapid, specific and convenient tool to detect simultaneously the Tobamoviruses.

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