• Title, Summary, Keyword: immunoperoxidase

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Modified Toluidine Blue: an Alternative Stain for Helicobacter pylori Detection in Routine Diagnostic Use and Post-eradication Confirmation for Gastric Cancer Prevention

  • Sakonlaya, Dussadee;Apisarnthanarak, Anucha;Yamada, Nobutaka;Tomtitchong, Prakitpunthu
    • Asian Pacific Journal of Cancer Prevention
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    • v.15 no.16
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    • pp.6983-6987
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    • 2014
  • Background: Modified toluidine blue staining (MTBs) is a simple, inexpensive and time saving method to detect H. pylori in gastric biopsy specimens. As a metachromatic stain, it simultaneously highlights intestinal metaplasia, a gastric cancer precancerous lesion. The aim of this study was to assess the reliability of MTBs compared with hematoxylin-eosin (H&E) for H. pylori detection using immunoperoxidase staining as the gold standard. This technique would be beneficial for a routine diagnosis and confirmation of H. pylori eradication in developing countries where endoscopic-based approaches are dominant. Materials and Methods: Esophagogastroduodenoscopy with triple site gastric biopsies was undertaken in 207 dyspeptic patients at Thammasat University Hospital, Thailand between 1997 and 1999. H&E, MTBs and immunoperoxidase staining were applied to each specimen. The presence or absence of H. pylori with each stain was interpreted separately and the sensitivity, specificity, positive and negative predictive values of H&E and MTBs were calculated. Results: A total of 282 specimens from 207 patients were evaluated. Using immunoperoxidase staining, organisms were positive in 117 specimens (41%). MTBs proved almost equally sensitive as immunoperoxidase (99%) and significantly more sensitive than H&E (85%). It has comparable specificity (96% vs 96%), PPV (95% vs 94%), and NPV (99% vs 90%) to H&E, using immunoperoxidase staining as gold standard. MTBs compared with immunoperoxidase staining, is cheaper (2 USD vs 12 USD) and faster (20 min vs 16 hrs) compared to immunoperoxidase staining. Conclusions: MTBs is effective, economical and easy to use in daily practice for the detection of H. pylori in gastric biopsy specimens. In addition to saving time in evaluating H. pylori associated gastritis, with a high sensitivity and ability to demonstrate intestinal metaplasia, the technique may have a role in confirmation of H. pylori eradication for gastric cancer prevention in a developing country setting.

Immunohistochemical identification of newcastle disease virus with indirect immunoperoxidase technique (Indirect Immunoperoxidase 법을 이용한 조직내 뉴켓슬병 바이러스 항원동정)

  • Nho, Whan-goog;Sur, Jung-hyang;Kim, Soon-bok
    • Korean Journal of Veterinary Research
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    • v.30 no.3
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    • pp.309-315
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    • 1990
  • The present experiment was done to identify newcastle disease virus(NDV) antigens in frozen sections of various oragns from experimentally NDV-infected with indirect immunoperoxidase method. Section were incubated with rabbit anti-NDV polyclonal as first antibody, followed by incubation with goat anti-rabbit or protein A peroxidase conjugate. Positive reactions were often detected in the epithelium of trachea and in the lymphocyte of spleen at 24 hours after virus inoculation. the viral antigen was localized mainly in the cytoplasm of infected cells. The method approved to be highly specific for the identification of NDV and allowed a precise localization of the viral antigens in infected cells.

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Immunohistochemistry for detection of Aujeszky's disease virus antigens : Immunogold-silver method in tissue sections (오제스키병 바이러스 항원검출을 위한 면역조직화학적 연구 -조직절편내 immunogold-silver법-)

  • Kim, Soon-bok
    • Korean Journal of Veterinary Research
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    • v.28 no.2
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    • pp.365-369
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    • 1988
  • The present study was done to demonstrate ADV antigens in frozen and paraffin sections from ADV-infected pigs and cell cultures by using of the IGS method. Tissue specimens from 3 young pigs infected with ADV-phylaxia strain and of 2 healthy pigs were used. Fibroblastic cells originated from pig brain and BHK cells were grown and confluent monolayers were infected with the virus. Two monoclonal antibodies and a specific hyperimmune serum to ADV were used as the source of primary antibodies for both the IGS and immunoperoxidase methods. Application of the IGS method yielded a black fine granular reaction in positive areas, and the results were superior to those obtained using the immunoperoxidase technique for all cases tested. The IGS method might be useful in the detection of various viral antigens in tissue sections.

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The Rapid Diagnosis of $\beta$-Haemolitic Streptococcus sp. by Immunoperoxidase Method (면역효소항체법에 의한 $\beta$-용혈성 연쇄구균 Streptococcus sp.의 신속진단에 대하여)

  • Chun, Seh-Kyu;Choi, Dong-Lim;Park, Soo-Il
    • Journal of fish pathology
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    • v.1 no.2
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    • pp.103-110
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    • 1988
  • For the rapid diagnosis of bacterial diseases of cultured fishes, the immunoperoxidase method was applied to the detection of $\beta$-haemolitic Streptococcus sp. strain KST-2 isolated from tilapia(Oreochromis niloticus). The suitability of field analysis and the sensitivity of the immunoperoxidase method was compared with those of the counterimmunoelectrophoresis(CIE) and the immunodiffusion(ID). Results of testing cross-reactivity which did not indicate any cross reactivity with other fish pathogens, this method was specific to Streptococcus sp. The sensitivity of this method was $1{\times}10^3CFU/ml$ which was at least $10^2$times greater than the CIE and $10^4$times greater than the ID. The immunoperoxidase method was more suitable for field application and more sensitive than other diagnostic techniques tested on this study.

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Detection of Retinol-binding Protein in Bovine Yolk Sac, Chorion and Allantois by Immunoperoxidase Method

  • Liu, Kaung Huei
    • Asian-Australasian Journal of Animal Sciences
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    • v.15 no.6
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    • pp.783-788
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    • 2002
  • Bovine yolk sac at day 24 of pregnancy, and placental membranes (chorion and allantois) from days 70 and 100 of pregnancy were isolated and cultured in a modified minimum essential medium in the presence of $[^{35}S]$methionine. Proteins synthesized and secreted by isolated bovine yolk sac, chorion and allantois were analyzed by fluorography of two-dimensional polyacrylamide gel electrophoresis. Serum-like proteins,transferrin, ${\alpha}$-fetoprotein, ${\alpha}$1-antitrypsin and ${\alpha}$1-acid glycoprotein,were the major protein products of yolk sac. A 21 kDa protein produced by yolk sac was identified immunochemically as retinol-binding protein (RBP). Chorion and allantios from days 70 and 100 of pregnancy were active in protein synthesis and secretion. Both chorion and allantois did not secret serum-like proteins but secreted a number of neutral-to-acidic proteins including RBP. Secretory proteins produced by the yolk sac, chorion and allantois may play important roles in the embryonic development and the successful outcome of pregnancy. Antiserum against bovine placental RBP was employed to the immunocytochemistry by immunoperoxidase method. Immunoreactive RBP was localized in epithelial cells and island-like cell clones of yolk sac. Immunostaining for RBP was detected in simple columnar epithelium of chorion and in simple squamous epithelium of allantois. In the present study, proteins synthesized and secreted by yolk sac at day 24 of pregnancy, chorion and allantois from days 70 and 100 of pregnancy were characterized In addition, RBP was localized in yolk sac, chorion and allantois by immunoperoxidase method. The immunoperoxidase method has been proven to be a very effective technique to identify the cellular source of protein synthesis in extraembryonic membranes.

Immunohistochemical diagnosis on rabies virus using its monoclonal antibody in mice (단크론항체를 이용한 광견병바이러스의 면역병리조직학적 진단)

  • Kang, Mun-il;Park, Nam-yong;Song, Jae-yeong
    • Korean Journal of Veterinary Research
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    • v.33 no.2
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    • pp.255-261
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    • 1993
  • For a immunohistochemical diagnosis of the frozen and paraffin-embedded tissues against rabies virus, mice were intracerebrally inoculated with challege virus standard(CVS) rabies virus and then were used to detect the rabies viral antigen by the immunoperoxidase(IP) and the avidin-biotin complex(ABC) method. In this study, the results confirmed that ABC and IP methods, although the former showed more specific and sensitive than the latter, were reliable and effective for the demonstration of rabies virus in both frozen and paraffin-embedded brain tissues prepared from rabies-infected mice. Additionally, IP technique using the monoclonal antibody against rabies virus could be recommended as a standard diagnostic tool instead of the present immunofluorescent method for the local veterinary services in Korea.

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Studies on the rabbit viral hepatitis : Immunohistochemical observations (토끼의 바이러스성 간염에 관한 연구 : 면역조직화학적 관찰)

  • Lee, Cha-soo;Shin, Tae-kyun;Choi, Youn-ju;Jeong, Kyu-sik;Jyeong, Jong-sik
    • Korean Journal of Veterinary Research
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    • v.33 no.2
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    • pp.269-275
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    • 1993
  • Tissue distribution of RHDV in rabbits were examined by immunofluorescence and ABC methods. Tissues including liver, spleen, kidneys, lungs and brain were frozen, cut in a crycut, and fixed in 10% buffered formalin, embedded in paraplast, and cut $5{\sim}7{\mu}m$ thickness. Sections were immunostained Tissue distribution of RHDV in rabbits were examined by immunofluorescence and ABC methods. Tissues including liver, spleen, kidneys, lungs and brain were frozen, cut in a crycut, and fixed in 10% buffered formalin, embedded in paraplast, and cut $5{\sim}7{\mu}m$ thickness. Sections were immunostained with primary antiserum and conjugated second antibodies as recommended by manufacturer. None of the cultures tested showed virus-induced phenomena. Immunoreactive products were commonly found in the liver, in some cases there were also positive staining in the spleen and kidneys. Other organs showed weak or insignificant immunoreactions. By ABC method on the formalin-fixed, paraffin-embedded liver tissues, strong immunoreactivity was found in the periportal triad lesions and peripheral lesions of the hepatic lobules. Immunoreactive products showed diffuse fine granular in the cytoplasm of hepatocytes and sinusoidal cells. In some cells, immunoproducts marginate at the periphery of the cells. The intensive staining of the cytoplasm of infected cells allowed their exact differentiation from surrounding uninfected cells. The positive area involved coincided with histopathological lesion on serial liver sections. In conclusion, liver was proved to be a consistent target organ in RHD, and the immunoperoxidase method in the section of formalin-fixed, paraffin-embedded hepatic tissue could be broadly used for the routine diagnosis of the disease.

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Immunohistochemical detection of viral antigen and pathological lesion in piglets experimentally infected with encephalomyocarditis virus (뇌심근염 바이러스의 실험적 감염자돈에 대한 병리학적 소견과 바이러스 항원의 면역조직화학적 검출)

  • Cho, Sung-hwna;Joo, Han-soo;Kim, Hyun-soo
    • Korean Journal of Veterinary Research
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    • v.33 no.2
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    • pp.301-308
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    • 1993
  • Three or 7day old piglets were infected experimentally with different encephalomyocarditis virus isolates to detect the viral antigen by the immunoperoxidase technique and to observe strain difference in their pathogenecity in newborn pigs by comparing clinical signs and pathologic lesions. Clinical signs of the infected pigs were different depending on the virus strain, pig age and infection route. Encephalomyocarditis virus(EMCV) NVSL-PR isolate was more pathogenic than MN-25 and MN-30 isolate. Three day old piglets showed more severe illness than 7 day old piglets. Predominant clinical signs were sudden death without noticeable clinical signs and dyspnea manifested as heavy abdominal breathing. Contact-infection from infected piglets to controls was observed in the oro-nasally infected group but not the intramuscular group. Common necropsy findings of dead piglets in both age groups infected with MN-25 and NVSL-PR were accumulation of excessive fluid in the body cavities and mild to diffuse necrotic areas observed in the hearts and occasionally in the livers. Microscopically, myocarditis with inflammatory cell infiltration, necrosis of the myocardial muscle fibers and occasional mineralization were observed along with interstitial pneumonia and centrolobular necrosis in the liver. Using an immunoperoxidase technique, viral antigen was detected in myocardial muscle fibers of piglets infected with EMCV.

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Production of monoclonal antibody(MAb) for lymphocytes of Korean native cattle 1. MAb to T lymphocytes (한우 림프구에 대한 단일항체 생산 1. T 림프구에 대한 단일항체)

  • Song, Hee-jong;Woon, Jae-ho;Kim, Tae-joong;Chai, Hyo-seok;Hur, Boo-hong;Choi, Min-soon
    • Korean Journal of Veterinary Research
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    • v.35 no.4
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    • pp.769-776
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    • 1995
  • This study was undertaken to develop the monoclonal antibody(MAb) for lymphocytes of Korean native cattle by the cell hybridization of myeloma P3/NS-1/ 1-Ag-4-1 and spleen cells from BALB/c mice hyperimmunized with nylon wool column eluted peripheral T lymphocytes of Korean native cattle. The isotype of MAb KCT-14 against T lymphocyte was mouse $IgG_1$. KCT-14 positivity of mononuclear cells(MNC) from peripheral blood lymphocytes, nylon wool nonadherent and adherent-lymphocytes was 41.7%, 58.4% and 22.6%, respectively. And that of mesenteric lymph node-, spleen and thymus-MNC was 43.3%, 40.2% and 33.6%, respectively. Immunoperoxidase staining of frozen tissue sections showed that the MAb positive cells were located in the medulla of the thymus and in the paracortical area and the mantle zone of the germinal center in the lymph nodes. These results indicated that KCT-14 was one of the MAb for investigate of T lymphocyte subpopulations in the Korean native cattle.

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Immunohistochemical study on the antigenicity of each organ structure of Clonorchis sinensis (간흡충 충체의 부위별 항원성에 대한 면역조직화학적 연구)

  • Jin Kim;Jong-Yil Chai;Weon-Gyu Kho;Kyu-Hyuk Cho;Soon-Hyung Lee
    • The Korean Journal of Parasitology
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    • v.29 no.1
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    • pp.21-30
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    • 1991
  • An immunohistochemical study was performed to demonstrate comparative antigenicity of each body structure of the liver fluke, Clonorchis sinensis, such as the digestive tract, reproductive organs, excretory system, tegument and suckers. Indirect immunoperoxidase technique was applied, rising formalin-fixed and paraffin-embedded sections of C. sinensis as the antigen. Pooled cat sera obtained 10 weeks after an experimental infection with C. sinensis and peroxidase-conjugated goat anti-cat IgG were used as the primary and secondary antibodies, respectively. The intensity of immunohistochemical stain was very sensitive upon the titers of the primary and secondary antibodies, and their optimum dilutions were found to be 1:1,000∼1:2,000 and 1:1,000, respectively. The intestinal epithelial cells, intestinal content and excretory bladder showed strong positive coloring reactions even at lower titer (1 : 2,000) of the primary antibody, whereas the uterine wall and eggs, vitelline glands, and male reproductive organs showed only weak positive reactions despite an increase in the antibody titer (1:1,000). On the other hand, the suckers, tegument, subtegumental cells and other parenchyme portions did not reveal any positive immunoperoxidase reaction at the same antibody titers. From the above results, it is highly suggested that the most potent antigenicity of C. sinensis occur from their excretory-secretory substances originated from the digestive and excretory organs.

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