• Title/Summary/Keyword: sandwich-ELISA

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Enzyme-Linked Immunosorbent Assay (ELISA) for Detection of Clostridium botulinum Type F Toxin (Clostridium botulinum Type F Toxin의 면역학적 효소방법에 의한 검출에 관한 연구)

  • Lee, Jeong-Kug;K. H. Yang
    • Microbiology and Biotechnology Letters
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    • v.10 no.3
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    • pp.205-209
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    • 1982
  • The enzyme-linked immunosorbent assay using the so-called "double-sandwich"technique was applied to determine Clostridium botulinum type F toxin. Polystyrene tubes were coated with horse anti-type F toxin serum and then toxin sample was added. The tubes were subsequently treated with rabbit anti-type F toxin IgG and sheep anti-rabbit serum IgG-horseradish peroxidase conjugate. By this technique, about 10 mouse intraperitoneal 50% lethal doses (ip LD/50/) of type F toxin could be detected. Low back-ground reading was achieved with the use of phosphate-buffered saline containing 0.05% Tween 20 and 1% bovine serum albumin as diluents of rabbit IgG and conjugate. Addition of EDTA in the diluents of toxin increased ELISA extinction value significantly. No cross-reaction was observed with botulinum type A and B toxin, but type E toxin gave sleight cross-reaction.

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Development and application of a monoclonal antibody-based sandwich ELISA for vitellogenin in carp (Cyprinus carpio)

  • Kang, Bong-Jung;Jeung, Jee-Hyun;Lee, Jae-Yong;Cho, Hyung-Koo;Kim, Myung-Hee;Han, Chang-Hee
    • Proceedings of the Korean Society of Fisheries Technology Conference
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    • 2003.05a
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    • pp.156-157
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    • 2003
  • In teleost, vitellogenin (Vtg) is synthesised in the liver in response to estrogens and transported by the blood to the growing oocytes where it is incorporated by micropincytosis (Selman and Wallace, 1982). Generally, Vtg is not induced in normal male fish but male fish are capable of synthesis Vtg in reponse to exogenous estrogen and xenoestrogens. (omitted)

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Detection of Cryptosporidium spp. in Diarrheic Immunocompetent Patients in Beni-Suef, Egypt: Insight into Epidemiology and Diagnosis

  • Gawad, Samah S. Abdel;Ismail, Mousa A.M.;Imam, Naglaa F.A.;Eassa, Ahmed H.A.;abu-Sarea, Enas Yahia
    • Parasites, Hosts and Diseases
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    • v.56 no.2
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    • pp.113-119
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    • 2018
  • Cryptosporidium species is an important cause of gastrointestinal infections globally. This study aimed to shed light on its role in diarrheic immunocompetent patients in Beni-Suef, Egypt and to compare three diagnostic methods. Two hundred diarrheic patients, $37{\pm}16.8year$ old, were enrolled. Stool samples were examined by light microscopy, using modified Ziehl-Neelsen stain (MZN) for Cryptosporidium spp. oocysts. Coproantigens were detected by sandwich ELISA. DNA molecular diagnosis was done by nested PCR. PCR yielded the highest detection rates (21.0%), compared to ELISA (12.5%) and MZN staining method (9.5%). The higher infection rates were in 20-40 year-old group, followed by 40-60 year-old. Association between epidemiologic factors was statistically not significant; positivity and gender, clinical manifestations, residence, source or water, or contact with animals. Cryptosporidiosis is an important enteric parasitic infection in Beni-Suef and PCR remains the gold standard for diagnosis.

Evaluation of ST2 and NT-proBNP as cardiac biomarkers in dogs with heartworm disease

  • Lee, Kiho;Park, Jun-Seok;Seo, Kyoung-Won;Song, Kun-Ho
    • Korean Journal of Veterinary Service
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    • v.41 no.2
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    • pp.79-83
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    • 2018
  • This study compared serum concentrations of suppression of tumorigenicity 2 (ST2) and N-terminal pro-B-type natriuretic peptide (NT-proBNP) between healthy and heartworm- infected dogs. Eighteen heartworm-infected dogs and five healthy dogs were included in the study. Dogs were diagnosed and categorized by history, clinical signs, and blood assay, thoracic radiography, echocardiography, and commercial ELISA kit results. Serum samples were sent to the IDEXX reference laboratory for NT-proBNP measurement. ST2 was examined by using a canine interleukin 33 receptor ELISA kit with the quantitative sandwich ELISA method. The severely infected group showed significant elevation of NT-proBNP concentration over those of the control (P=0.03) and mildly infected (P=0.04) group. There were no significant difference in ST2 concentrations among the three groups. The usefulness of NT-proBNP as a cardiac biomarker in dogs with severe heartworm disease was confirmed by the results of this study. Further investigations to assess ST2 as a cardiac biomarker are warranted.

Detecting Activated Thrombin Activatable Fibrinolysis Inhibitor (TAFIa) and Inactivated TAFIa (TAFIai) in Normal and Hemophilia A Plasmas

  • Hulme, John P.;An, Seong Soo A.
    • Bulletin of the Korean Chemical Society
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    • v.30 no.1
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    • pp.77-82
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    • 2009
  • Thrombin activatable fibrinolysis inhibitor (TAFI) also known as plasma procarboxypeptidase B or U is a 60 kD glycoprotein, which is the major modulator of fibrinolysis in plasma. TAFI is a proenzyme, which is activated by proteolytic cleavage to an active carboxypeptidase B-like enzyme (TAFIa, 35.8 kD) by thrombin/thrombomodulin and plasmin. Modulation of fibrinolysis occurs when TAFIa enzymatically removes C-terminal lysine residues of partially degraded fibrin, thereby inhibiting the stimulation of tissue plasminogen activator (t-PA) modulated plasminogen activation. TAFIa undergoes a rapid conformational change at $37{^{\circ}C}$ to an inactive isoform called TAFIai. Potato tuber carboxypetidase inhibitor (PTCI) was shown to specifically bind to TAFIa as well as TAFIai. In this study, a novel immunoassay TAFIa/ai ELISA was used for quantitation of the two TAFI activation isoforms TAFIa and TAFIai. The ELISA utilizes PTCI as the capture agent and a double antibody sandwich technique for the detection. Low levels of TAFIa/ai antigen levels were detected in normal plasma and elevated levels were found in hemophilia A plasmas. TAFIa/ai antigen represents a novel marker to monitor fibrinolysis and TAFIa/ai ELISA may be a valuable assay for studying the role of TAFI in normal hemostasis and in pathological conditions.

A Study on Serum Leptin Values by Elisa Method in Children (ELISA법으로 측정한 소아 혈중 LEPTIN 치에 관한 연구)

  • Song, Soo-Ho;Chung, Young-Hun
    • Pediatric Gastroenterology, Hepatology & Nutrition
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    • v.3 no.2
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    • pp.175-180
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    • 2000
  • Purpose: Leptin is an adipocyte-derived blood-borne satiety factor that acts on its cognate leptin receptor in the hypothalamus, thereby regulating food intake and energy expenditure. We measured the leptin concentrations in serum of normal and obese children with human leptin ELISA kit, unlike previous study with leptin RIA kit and investigated the relationship between leptin concentrations and body mass index, gender, and age. Methods: We measured serum concentrations of leptin in 67 children who were visited to the Department of Pediatrics, Chungnam National University Hospital during the period of 5 months from February, 1999 to June, 1999. Height, weight, obesity index, and body mass index were measured in 67 subjects. Leptin values in serum were measured by sandwich ELISA method. Data analysis was done according to the obesity, body mass index, gender and age. Results: The mean concentration of leptin was $7.69{\pm}8.83\;ng/ml$ in normal children group and $36.34{\pm}18.57\;ng/ml$ in obese group. Serum leptin concentrations were significant correlation with the body mass index (p<0.01). Serum leptin concentration was significant higher in the group of over 10 years of age (p<0.01). Leptin levels showed no significant difference by gender. Conclusion: Serum leptin levels were significantly higher in obesity group than in control one, and they were correlated with body mass index and age. Measurements of leptin value by sandwich ELISA method are very useful and easily applicable to determine obesity.

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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.

The Changes of Allergenic and Antigenic Properties of Major Allergen(Pen a 1) of Brown Shrimp(Penaeus aztecus) by Gamma Irradiation (감마선 조사에 의한 갈색 새우(Penaeus aztecus) 주요알러젠(Pen a 1)의 알러지성 및 항원성의 변화)

  • Lee, Ju-Woon;Kim, Jae-Hun;Sung, Chang-Keun;Kang, Kun-Ok;Shin, Myung-Gon;Byun, Myung-Woo
    • Korean Journal of Food Science and Technology
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    • v.32 no.4
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    • pp.822-827
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    • 2000
  • Gamma irradiation was applied to reduce shrimp allergy. Shrimp heat-stable protein(HSP) and shrimp protein extract were gamma-irradiated at 1, 3, 5, 7 or 10 kGy in an aqueous state (1.0 mg/mL). The changes in allergenic and antigenic properties of protein extract and HSP resulted from gamma irradiation were monitored by ELISA with mouse mAb or human patients sera and immunoblotting. Conformational changes in irradiated HSP were measured by both GPC-HPLC and SDS-PAGE. The binding ability of shrimp allergic patients IgE to irradiated protein extract or irradiated heat-stable protein was dose-dependently reduced. When measured by gel permeation chromatography and sandwich ELISA, the amount of intact heat-stable protein in the irradiated solution was reduced by gamma irradiation depending upon the applied dose. SDS-PAGE showed that the main band disappeared and new bands appeared in a higher molecular weight zone. The results provide a new possibility to use irradiation process for reducing the allergenicity of shrimp.

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Increased Interlenkin-2 Serum Level in Male Schizophrenic Patients (남자 정신분열증 환자에서 혈청 Interlenkin-2 농도의 증가)

  • Kim, Yong-Ku;Kim, Sa-Jun;Lee, Min-Soo
    • Korean Journal of Biological Psychiatry
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    • v.3 no.1
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    • pp.109-114
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    • 1996
  • We have previously reported that Korean schizophrenic patients hove low production of IL-2 in vitro suggestive of autoimmunity to the pathogenesis of the disorder. In an attempt to further explore this issue, we measured in vivo serum levels of interleukins(IL-$1{\beta}$, IL-2, and IL-6) using a quantitative "sandwich" enzyme immunoassay(ELISA) in 26 male schizophrenic patients and in 26 age-matched normal controls. Patients met DSM-IV criteria for schizophrenia and were drug free for at least six months. The severity of symptoms was assessed by SANS and SAPS. We found a significant increase of IL-2 level(p<0.05) in schizophrenic patients as compared with normal controls. There were significant positive correlations between IL-2, IL-6 levels and negative symptom scores. There were no correlations between age, age at onset, duration of illness and interleukin levels. Our results may support the hypothesis of viral-autoimmune dysfunction in schizophrenia. IL-2 or IL-6 may be associated with specific clinical feature in schizophrenic syndrome, especially negative symptom.

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