• Title/Summary/Keyword: ELISA antibody

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A Recombinant $Plasmodium$ $vivax$ Apical Membrane Antigen-1 to Detect Human Infection in Iran

  • Haghi, Afsaneh Motevalli;Khoramizade, Mohammad Reza;Nateghpour, Mehdi;Mohebali, Mehdi;Edrissian, Gholam Hossein;Eshraghian, Mohammad Reza;Sepehrizadeh, Zargham
    • Parasites, Hosts and Diseases
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    • v.50 no.1
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    • pp.15-21
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    • 2012
  • In Iran, $Plasmodium$ $vivax$ is responsible for more than 80% of the infected cases of malaria per year. Control interventions for vivax malaria in humans rely mainly on developed diagnostic methods. Recombinant $P.$ $vivax$ apical membrane antigen-1 (rPvAMA-1) has been reported to achieve designing rapid, sensitive, and specific molecular diagnosis. This study aimed to perform isolation and expression of a rPvAMA-1, derived from Iranian patients residing in an endemic area. Then, the diagnostic efficiency of the characterized Iranian PvAMA-1 was assessed using an indirect ELISA method. For this purpose, a partial region of AMA-1 gene was amplified, cloned, and expressed in pET32a plasmid. The recombinant $His-tagged$ protein was purified and used to coat the ELISA plate. Antibody detection was assessed by indirect ELISA using rPvAMA-1. The validity of the ELISA method for detection of anti-$P.$ $vivax$ antibodies in the field was compared to light microscopy on 84 confirmed $P.$ $vivax$ patients and compared to 84 non-$P.$ $vivax$ infected individuals. The ELISA cut-off value was calculated as the mean+2SD of OD values of the people living in malaria endemic areas from a south part of Iran. We found a cut-off point of OD=0.311 that showed the best correlation between the sera confirmed with $P.$ $vivax$ infection and healthy control sera. A sensitivity of 81.0% and specificity of 84.5% were found at this cut off titer. A good degree of statistical agreement was found between ELISA using rPvAMA-1 and light microscopy (0.827) by Kappa analysis.

Serodiagnosis of cysticercosis by ELISA-inhibition test using monoclonal antibodies (단세포군항체를 이용한 효소면역억제측정 법에 의한 유구낭미충증의 혈청학적 진단)

  • 용태순;여인석
    • Parasites, Hosts and Diseases
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    • v.31 no.2
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    • pp.149-156
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    • 1993
  • Monoclonal antibodies (Mabs) were produced against crude scolex extract of T solium metacestodes, and applied to ELISA-inhibition test for improving the specificity of serodiagnosis of human cysticercosis. Four hybridomas secreting species-specific anti- cysticercal Mabs (Cya-1, Cya-7, Cya-28 and Cya-31) were selected. Each Mab reacted on antigenic components of 25.5 kDa (Cya-1), 28 kDa (rcya-7), 87.5 kDa (Cya-281), and 12.5 kDa (Cya-31). IFA showed that Cya-1 was located at the calcium corpuscles, and Cya-7 at the loose connective tissue of T soLium metacestode scolex. Cya-28 and Cya-31 reacted on the tegument of the scolex. By conventional ELISA, 23 out of 28 (82.1%) cysticercosis patients were found serologically positive, but 1 out of 9 (11.1%) sparganosls cases and 6 out of 31 (19.4%) paragonlmiasis cases showed false positives. By ELISA-Inhibition test using species-specific anti-cysticercal Mab Cya-7, 19 out of 28 (67.9%) cystlcercosis cases were found serologically positive, but there were no false positives In other parasitic infections.

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Porcine Circovirus Infection in Weaned Pigs with Postweaning Multisystemic Wasting Syndrome in Korea (국내 이유자돈의 써코바이러스 감염에 의한 이유후전신소모성 증후군)

  • Kim, Jae-hoon;Roh, In-soon;Sohn, Hyun-joo;Jean, Young-hwa;Hwang, Eui-kyung;Yoon, Kyoung-jin
    • Korean Journal of Veterinary Research
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    • v.43 no.3
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    • pp.463-469
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    • 2003
  • Eight nursery to grower pigs exhibiting weight loss and sudden death were diagnosed as postweaning multisystemic wasting syndrome (PMWS) based on the results of gross findings, histopathology, immunohistochemistry, fluorescent antibody test, virus isolation, PCR, serology, and electron microscopy. Groosly, the pigs had a rough hair coats and were severely emaciated. And moot lymph nodes were pale and enlarged. Lungs were not fully collapsed and exhibited 10 to 40% pale red cranioventral consolidation. Histopathologically, typical lymphohistiocytic interstitial to bronchointerstitial pneumonia, chronic lymphadenitis, severe lymphoid depletion, and basophilic intracytoplasmic inclusions were noted in the most lymphoid tissues. Porcine circovirus panicles were observed in the inguinal lymph node of the pigs by electron microscopy. Porcine circovirus type 2 (PCV2) antigens or viral DNAs were detected in the lesions of all pigs using immunohistochemistry or PCR. Two PCV2 were isolated from a homogenate of pooled lung and lymph node in 2 of the 5 pigs. Additionally, antigens of porcine reproductive and respiratory syndrome virus (PRRSV) and Hemophilus (H.) parasuis were also detected by immunofluorescent antibody test. Serologically, 55% of randomly selected sows and fattening pigs was serum antibody positive to PCV2 by an indirect fluorescent antibody (IFA) test and approximately 18 % of animals in the herd were serologically pooitive by the ELISA kit for PRRSV. To our knowledge, this is the first report of PMWS co-infected with PCV-2, PRRS, and H. parasuis in Korea.

Field efficacy of a combined vaccine supplemented with recombinant Pasteurella multocida toxin subunits against atrophic rhinitis

  • Kang, Mi Lan;Shin, Seung Won;Rayamahji, Nabin;Seo, Yeon Soo;Lee, Su In;Lee, Won Hyung;Yoo, Han Sang
    • Korean Journal of Veterinary Research
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    • v.48 no.1
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    • pp.53-60
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    • 2008
  • We have investigated efficiency of a recombinant subunit Pasteurella multocida toxin (PMT) that was mixed with a vaccine consisted of inactivated whole cells of Bordetella bronchiseptica, P. multocida (types A and D). For verification of the efficacy of the vaccine, all experimental pigs (suckling piglets, sow and gilts) in the three farms were vaccinated. Antibody titers against B. bronchiseptica and P. multocida type A of the vaccinated pigs by microplate agglutination were significantly higher than those of the control pigs (p < 0.05). Similar patterns were observed in the analysis of anti- PMT neutralizing antibody by serum neutralizing method using Vero cell (p < 0.05). Anti- P. multocida type D antibody titer of the vaccinated sows and gilts by ELISA showed significant differences with those of the non-vaccinated pigs (p < 0.05). Although antibody titers increased, it was unable to find out the difference in the clinical signs between the vaccinated and non-vaccinated pigs. However, the increase in body weight of the vaccinated piglets was observed in comparison with the non-vaccinated piglets on a farm. At slaughtering of the pigs, pathological lesions in the turbinate bones of the vaccinated pigs were significantly lower than those of the non-vaccinated pigs (p < 0.001). These results suggested that efficacy of the vaccine in pigs demonstrated to protect against atrophic rhinitis in Korea.

Sero-prevalence of swine influenza virus (H1N1, H3N2) In Korea (국내의 돼지 인플루엔자 바이러스(H1N1, H3N2)의 혈청학적 조사)

  • Yoon, Jai-soon;Park, Bong-kyun;Han, Jeong-hee
    • Korean Journal of Veterinary Research
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    • v.47 no.3
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    • pp.273-279
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    • 2007
  • Swine influenza is an acute, infectious respiratory disease caused by type A influenza viruses in pigs. In the previous studies, serological surveys have indicated the presence of H3N2 swine influenza virus (SIV) since 1995 in Korea. And the percentage of the antibody-positive rate was 39.12% in the survey determining the prevalence of H1N1 SIV antibodies in 2002. The purpose of this study was therefore to investigate the sero-prevalence of SIV regard to the age of the pig and the season between June 2004 and May 2005. In this study, a total of 932 sera were used. These sera were randomly selected from blood samples, which were submitted to Department of Veterinary Pathology, Kangwon National University and Department of Veterinary Virology, Seoul National University from June 2004 to May 2005. These sera have been tested by ELISA test kit (IDEXX Lab, USA) for the SIV H3N2, H1N1 respectively. SAS version 9.1 was used for the statistical analysis based on the age of the pig and the season. The overall sero-prevalence of the antibody against H3N2 SIV was 20.82% (194/932). The overall sero-prevalence of the antibody against H1N1 SIV was 37.23% (347/932). The overall dual sero-prevalence of the antibody against H3N2 and H1N1 SIV was 10.62% (99/932). H3N2 has significant difference in statistically regarding the age of the pig and the season (p<0.0001). H1N1 has significant difference in statistically regarding the age of the pig (p<0.0001) but has not significant difference in statistically regarding the season (p=0.5882).

Production of Monoclonal Antibody to Polychlorinated Biphenyl Induced Cytochrome P-450 LMII in Rat Liver (Polychlorinated Biphenyl에 의한 백서간 Cytochrome P-$450_{LMII}$에 대한 Monoclonal Antibody 생성에 관한 연구)

  • Kim, Jung-Hye;Kim, Jae-Ryong;Lee, Ki-Yung
    • Journal of Yeungnam Medical Science
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    • v.3 no.1
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    • pp.103-110
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    • 1986
  • Cytochrome P-450(CP-450) is one of the three components of the liver microsomal enzyme system which hydroxylates fatty acids, hydrocarbons and a variety of drugs and other foreign compounds. Female Balb/c mice were immunized with purified polychlorinated bipheny(PCB)-induced CP-450 LMII. The spleen cells derived from immunized mice were fused with $SP^2$ myeloma cells using polyethylene glycol(PEG 3500). The hybrid cells were selected by hypoxanthine-aminopterine and thymidine(HAT) medium and the culture fluid were screened by enzyme-linked immunosorbent assay to CP450 LMII. The hybrid cess(${\times}10^7$) were innoculated into intraperitoneal cavity of Balb/c mice for the purpose of production of ascitic fluids. Monoclonal antibody(Mab) was purified from ascitic fluid by DEAE cellulose ion exchange chromatography and $I^{125}$-labeled Mab was also confirmed by autoradiography and SDS-polyacrylamide gel electrophoresis (MW : 55,000 and 110,000).

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The Production of Egg Yolk Immurnoglobulin (IgY) Raised against 3T3L-1 Cell Membrane Protein and the Control of Adipocytes Differentiation (3T3L-1세포의 막단백질에 대한 난황면역글로뷸린 (IgY)의 생산과 지방세포의 분화조절작용)

  • 김상윤;황성구;구의섭;고태송
    • Korean Journal of Poultry Science
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    • v.26 no.3
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    • pp.179-188
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    • 1999
  • The present was undertaken to establish a model for the control of adipocytes differentiation by using antibody from egg yolk. The emulsion of membrane protein of 3T3L-1 cell membrane protein with the complete Freund's adjuvant was firstly immunized in layer. Second and third boosting were undertaken with two weeks intervals by injection of the emulsion of the same antigen with the incomplete Freund's adjuvant. After 4 week of the first immunization, eggs were collected and antibody (IgY) was purified from egg yolk. The purity of IgY was 60-98% determined by single radial immunodiffusion (SRID) methods. Titer value of the antibody showed high reactiviy for the preadipocytes membrane protein measured by ELISA. When the IgY was added in the test media containing either 2.5% porcine serum or 10% FBS(control), the differentiation of 3T3L-1 cells and Glycerol-3-phosphate dehydrogenase(GPDH) activities was significantly decreased compared to the control cells(p〈0.05). When mice were subcutaneously injected with IgY raised against membrane protein of 3T3L-1 cells for 3 weeks, adipose tissue mass around ovary was tended to be decreased in female mice compared to those of control mice. It is suggested that a potential for manipulating of lipid accumulation through decrease in 3T3L-1 cell differentiation and fat accumulation in female mice by IgY treatment.

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Immunoblot patterns of clonorchiasis (면역이적법에 의한 간흡충 항원분획과 감염자의 항체반응 양상)

  • 홍성태;고원규
    • Parasites, Hosts and Diseases
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    • v.35 no.2
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    • pp.87-94
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    • 1997
  • Clonorchis sinearis is a liver fluke which is the most prevalent helminth of humans in Korea. The better diagnostic measure of clonorchiasis is required for its nationwide control program. The present study observed antigenic bands of C. sinensis and reacting immunoglobulins in serum of infected residents. Adult C. sinensis were recovered from experimentally infected rabbits and soluble crude extract of the worms was used as the antigen after supplementation of E-64, a cysteine proteinase inhibitor. SDS- PAGE of the crude antigen resolved more than 20 protein bands between 200 and 14 kDa. The sera of infected humans collected at an endemic village showed specific IgG and IgE antibodies but little IgM and IgA antibodies. The protein bands of 94, 80, 72, 68, 52, 47, 43, 37, 34, and 28-25 kDa strongly reacted with serum Ig(GMA) or IgG antibody and 64, 62, 52, 47,44, 34,28, and 26 kDa bands reacted with serum IgE antibody. However, the 94, 80, 72, 68, 64, 62, 52, 47, and 40 kDa bands of C. sinensis antigen were found non specific. The protein bands of 43, 34, and 28-25 kDa of C. sinensis are primary target molecules of further analysis.

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Effects of infectious bursal disease virus(IBDV) and newcastle disease virus(NDV) vaccines on performance of broiler chicks

  • Kwon, Jung-taek;Kim, Tae-joong;Ryu, Kyeong-seon;Song, Hee-jong
    • Korean Journal of Veterinary Research
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    • v.39 no.4
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    • pp.738-742
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    • 1999
  • The objective of this experiment was to investigate the effect of Newcastle disease virus (NDV) and infectious bursal disease virus (IBDV) vaccination on performance of broiler chicks for five weeks. Two types of poultry houses and three patterns of vaccination ($NDV^-/IBDV^-$, $NDV^+/IBDV^-$ and $NDV^+/IBDV^+$) were factorially assigned to six treatments. NDV, B1 strain and IBDV, Bursin-2 vaccine were orally administered at 5, 14 and 7, 18 days, respectively. Forty eight hundred chicks were grouped into four replications with two hundnyd hybro $\times$ hybro chicks per each treatment. Weight gain, feed conversion ratio (FCR), mortality and product index were surveyed at the end of experiment. Bursa index and IBDV antibody titer of chicks were weekly measured. Weight gain of chicks vaccinated with $NDV^+/IBDV^+$ was significantly increased compared to that of other treatments at both window and windowless poultry houses (p<0.05). Chicks vaccinated with $NDV^+/IBDV^+$ also showed significantly improving the FCR and mortality compared to those of other treatments at both poultry houses (p<0.05). The bursa indecies of both poultry houses were high from one-day- to three-weeks-old, but were low for the rest of two weeks. IBDV antibody of all chicks was detected 100% by agar gel precipitation (AGP) test at one day old, but was not detected in $NDV^-/IBDV^-$ and $NDV^+/IBDV^-$ treatments at four weeks old. However, it showed 100% in $NDV^+/IBDV^+$ treatment. Antibody titer using ELISA showed similar trend to that of AGP test. The results of this experiment confirmed that IBDV and NDV combined vaccine significantly improved the performance of broiler chicks.

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Production of polyclonal anti-$\beta$-adrenergic receptor antibody and it′s effects on receptor ligand binding

  • Kim, Hee-Jin;Shin, Chan-Young;Noh, Min-Su;Ko, Kwang-Ho
    • Proceedings of the Korean Society of Applied Pharmacology
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    • 1995.04a
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    • pp.86-86
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    • 1995
  • The analysis of membrane receptors for hormones and neurotransmitters has progressed considerably by pharmacological and biochemical means and more recently by the use of specific anti-receptor antibodies. A 14-mer peptide (from Phe102 to Leu115 of ${\beta}$2-adrenergic receptor) was synthesized and this peptide was coupled to carrier protein Keyhole Limpet Hemocyanin(KLH) by glutaraldehyde method. A 0.5mg of KLH-coupled peptide was emulsified with equal volume of complete Freund's adjuvant and injected via popliteal lymph node to each of the three Newzealnd White rabbits. Booster injections were repeated at 4 weeks interval for three times with incomplete Freund's adjuvants. One week after the final injection, serum was prepared from ear artery. Nonspecific immunoglobulins were removed by passing the serum through KLH-Sepharose 6B affinity matrix and further by incubation with bovine lung aceton powder. The titer of the antibody for synthetic peptide which was determined by enzyme linked immunosorbent assay(ELISA) was about l/l,000. The antibody produced in this study revealed 67kDa protein band in the western blot of partially purified guinea pig lung ${\beta}$-adrenergic receptor preparation. The antibody inhibited ${\beta}$-adrenergic antaginist [3H] Dihydroalprenolol binding to soluble ${\beta}$-adrenergic receptor by 25% while control sera did not show any inhibitory effects, The result of this study suggests that the peptide sequence selected in this study may play some important roles in adrenergic receptor-ligand interaction.

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