• Title/Summary/Keyword: Immunogold labeling

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Immunoelectron Microscopic Localization and Analysis of Herpes simplex Virus Type 1 Antigens

  • Chung, Charles C.;Lee, Hyung-Hoan;Cho, Myung-Hwan
    • Journal of Microbiology and Biotechnology
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    • v.10 no.5
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    • pp.714-720
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    • 2000
  • Antigens of Herpes simplex virus type 1 (HSV-1) strain F were immunoblotted to identify the most immunodominant one, and the localization of this antigen was then studied using immunoelectron microscopy. The 67.8 kDa antigen appeared to be the most immunodominant one in a mouse model, and it showed randomly scattered and partially clustered distribution on the surface of the virion. The localization study was performed using immunogold with polyclonal anti-HSV-1 sera produced from BALB/c mice, and immunofluorescence demonstrated that the viral products in the HSV-2 infected Vero cells were distributed throughout the infected host cell, however, mainly on the surface of the host membrane.

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A study on the body fluid antigen of Clonorchis sinensis using immunogold labeling method (면역황금 표기법을 이용한 간흡충의 체액 항원에 관한 연구)

  • Ju, Bong-Deok;Im, Han-Jong;Kim, Su-Jin
    • Parasites, Hosts and Diseases
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    • v.28 no.1
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    • pp.11-24
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    • 1990
  • In order to observe the antigenic localization in the tissues of the adult Clonorchis sinensis, immunogold labeling method was applied using serum immunoglobulins (IgG) of either worm·infected rabbits (group I) or antigen-immunized rabbits (group II) (by the body quid obtained from the adult worms). The electron micrographs of the sectioned worm tissue antigens, embedded in Lowicryl HM 20 medium and stained with protein A-gold complex (particle sixte: 12 nm), were compared between the group I and group II. The gold particles were observed in the interstitial matrix of the worm parenchyma, the epithelial lamellae of the cecum, and the cecal lumen both in group I and II. But the particles were in general more concentrated in group II. The gold particles were not observed on the basal lamina of the tegument or on vitelline glands in group I, while they were highly concentrated on those areas in group II. There were also differences in the antigenicity of interstitial matrix(reacted with group I IgG) and head part(reacted with group II IgG) of the sperm cells in the seminal receptacle. Conclusively, it is suggested that the substances comprising the basal lamina of the tegument or vitelline glands act as specific antigens reacting with antigen(body quid) immunized rabbit IgG. On the other hand, the substances in the cecal lumen and cecal epithelial lamellae are thought to be the specific antigen that react with the worm-infected rabbit IgG.

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Immuno-Electron Microscopic Studies on the Distribution of Dopamine and $Calbindin-D_{28K}$ in the Optic lobes of Cephalopods (Todarodes pacificus and Octopus minor) inhabiting the Korean waters (한국 연근해산 두족류 (Todarodes pacificus and Octopus minor) 시엽내 Dopamine 및 $Calbindin-D_{28K}$의 분포에 관한 면역전자현미경적 연구)

  • Han, Jong-Min;Chang, Nam-Sub
    • Applied Microscopy
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    • v.32 no.2
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    • pp.175-183
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    • 2002
  • In this study, we carried out immunostaining and immunogold labeling with rabbit anti-dopamine (TH) and rabbit anti-calbindin-$D_{28K}$ to examine the characteristics and functions of the neurons that secrete neurotransmitters in optic lobes of Todarodes pacificus and Octopus minor inhabiting the Korean waters. The obtained results are as follow. In the immunostaining with anti-dopamine, only a few of the large amacrine cells in an the upper part of an outer granule cell layer and the cells forming the islands of medulla showed positive reaction in Todarodes pacificus, while $2{\sim}3$ cells in the upper and middle parts of an outer granule cell layer and more than 5 cells in the islands of medulla reacted positively in Octopus minor. For the case of anti-calbindin case, $2{\sim}3$ small amacrine cells in the upper portion of the outer granule cell layer and $1{\sim}2$ cells which are located in the lower part of an inner granule cell layer showed positive reaction in Todarodes pacificus, while, in Octopus minor, 4 cells in the outer granule cell layer reacted positively, no immunoreactive cell being found in the inner granule cell layer. As a result of performing the immunogold labeling, relative large number ($17{\sim}26$) of gold particles were labeled per $0.5{\mu}m^2$ of the cytoplasm of the cells which showed the immunoreactivity to the anti-dopamine and anti-calbindin in Todarodes pacificus, however, small number (10) of gold particles were labeled in Octopus minor, which reach only half of the number in the Todarodes pacificus.

Ultrastructural Localization of Cryptosporidium parvum Antigen Using Human Patients Sera

  • Lee, Jong-Gyu;Han, Eun-Taek;Park, Woo-Yoon;Yu, Jae-Ran
    • Parasites, Hosts and Diseases
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    • v.47 no.2
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    • pp.171-174
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    • 2009
  • The antigen location of Cryptosporidium parvum, which stimulates antibody formation in humans and animals, was investigated using infected human sera. Immuno-electron microscopy revealed that antigenicity-inducing humoral immunity was located at various developmental stages of parasites, including asexual, sexual stages, and oocysts. The amount of antigen-stimulating IgG antibodies was particularly high on the oocyst wall. The sporozoite surface was shown to give stimulation on IgG and IgM antibody formation. Trophozoites implicated the lowest antigenicity to humoral immunity, both IgG and IgM, by showing the least amount of gold labeling. Immunogold labeling also provided clues that antigens were presented to the host-cell cytoplasm via feeder organelles and host-parasite junctions.

Application on Microwave Energy in the Preparation of Fish Samples for Electron Microscopic Observation

  • Kim Soo Jin;Oh Hae Keun;Song Young-Hwan;Chung Hyun-Do;Kim Young-Tae;Park Nam-Kyu;Choi Tae-Jin
    • Fisheries and Aquatic Sciences
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    • v.1 no.2
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    • pp.187-191
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    • 1998
  • Chemotherapy can not be applied for the control of fish viral diseases because viruses depend on host machinery for their replication. Although new control strategies including vaccination are under development, avoidance of virus introduction by rapid and correct diagnosis is the best way of fish viral disease control. Although observation of virus particles with an electron microscope is an easy method for virus detection, it take a few days for the sample preparation. In order to shorten the sample preparation time, microwave radiation was applied in the procedure. With this method, 15 seconds was enough for fixation of virus infected fish samples or cultured cells inoculated with infectious hematopoietic necrosis virus, which takes 2-4 hours with routine methods. Also four minutes was enough for polymerization of embedding resin which takes 24-48 hours with routine methods. Samples prepared with microwave were good enough for direct electron microscopic observation and immunogold labeling assay.

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Distribution of Hemicelluloses in Warts and the Warty Layer in Normal and Compression Wood Tracheids of Cryptomeria Japonica

  • Kim, Jong-Sik;Awano, Tatsuya;Yoshinaga, Arata;Takabe, Keiji
    • Journal of the Korean Wood Science and Technology
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    • v.39 no.5
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    • pp.420-428
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    • 2011
  • The distribution of arabino-4-O-methylglucuronoxylans (AGXs) and O-acetyl-galactoglucomannans (GGMs) in warts and the warty layer of tracheids in normal wood (NW) and compression wood (CW) of Cryptomeria japonica was investigated. Under field emission scanning electron microscope (FE-SEM) observation, warts and the warty layer of delignified NW and CW tracheids were degraded by xylanase treatment, indicating that warts and the warty layer contain high amounts of AGXs. However, the effect of xylanase was not observed in NW and CW tracheids before delignification, suggesting that AGXs in warts and the warty layer may be encrusted with lignin. After ${\beta}$-mannanase treatment, no noticeable changes were observed in warts and the warty layer of NW tracheids, indicating that warts and the warty layer contain either no or very few GGMs. Similar results to FE-SEM observations were also observed with immunogold labeling. AGX labeling was observed in warts and the warty layer of NW and CW tracheids, while GGM labeling was not detected. NW tracheids showed a much stronger density of AGX labeling than did CW tracheids in warts and the warty layer, indicating differences in the chemical compositions of warts and the warty layer between NW and CW tracheids.

Localization of Single Chain Fv Antibodies (scFv) in Transgenic Tobacco Ptants Showing Resistance against Tomato Bushy Stunt Virus

  • Jeun, Y.C.;Boonrod, K.;Nagy, P.;Conrad, U.;Krczal, G.
    • Proceedings of the Korean Society of Plant Pathology Conference
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    • 2003.10a
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    • pp.75.2-75
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    • 2003
  • To develop an effective protection strategy against tomato bushy stunt virus (TBSV), tobacco plants expressing single-chain Fv antibodies (scFv), were established. A previous had shown that the replication activity of viral replicase was inhibited by the selected scFvs. Moreover, no systemic symptom was found after virus inoculation on leaves of wt N. benthamiana infiltrated with an Agrobacterium suspension resulting i3l expression of the scFvs. However, control plants showed systemic symptoms. In this study the localization of the scFvs within two transgenic plant lines, (CP28H3, CP-P55) was demonstrated using immunogold labelling. The gold particles, indicating the presence of scFv, were mostly found In the cytoplasm of the plant cells including chloroplasts and in the cell walls. However, they were hardly found in the vacuole, nucleoplasm and intercellular spaces. Gold particles often accumulated in either the cytosol or chloroplasts showing a specific labeling, There was no difference in type of gold labeling between both transgenic lines. The localization of the scFv in the cytoplasm further conforms the inhibition of the RNA-dependent RNA polymerase (RdRp) by the selected scFv because it is known that the RdRp is localized to membraneous cytosolic structures.

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A Study on the Distribution of Cytochrome-c-oxidase Subunit in the Cristae of Mitochondria (미토콘드리아 크리스테에 존재하는 cytochrome-c-oxidase의 단백질 소단위 분포에 관한 연구)

  • Kim, Soo-Jin;Lee, Ji-Hyon;Chung, Cha-Kwon
    • Applied Microscopy
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    • v.24 no.4
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    • pp.41-51
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    • 1994
  • The topology of the enzyme has been investigated by biochemical studies including chemical labeling and cross linking. Thirteen subunits(polypeptides) of the cytochrome-c-oxidase have localistic characteristics of existing in the matrix side or cytoplasmic side in the mitochondria. In order to observe the distribution of the enzyme subunit on the mitochondria membrane, immunogold-labeling methods were employed. Antibody was obtained from the serum of immunized rabbit with enzyme subunit antigen which was obtained from cytochrome-c-oxidase of the beef heart muscle mitochondria. Beef heart muscle tissue as a tissue antigen was stained with immunized rabbit IgG and protein A gold complex. Electron microscopy has identified the existance of cytochrome-c-oxidase subunit $Mt_I,\;Mt_{II}\;and\;Mt_{III}$ on the membrane of cristae and outer chamber of mitochondria and the subunit $C_{IV}$ on the membrane of cristae and matrix of mitochondria. Particularly, the subunit $C_{IV}$ was also observed to exist in the sarcoplasm of muscle tissue.

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Binding of Tp92 homolog of Treponema denticola to fibronectin and epithelial cells

  • Jun, Hye-Kyoung;Lee, Sung-Hoon;Lee, Hae-Ri;Choi, Bong-Kyu
    • International Journal of Oral Biology
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    • v.33 no.2
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    • pp.45-50
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    • 2008
  • Treponema denticola is the best studied oral spirochete and numerous studies have shown that it is strongly associated with periodontitis and expresses several putative virulence factors. In this study, we report on a surface protein of T. denticola, Td92, which is homologous to Tp92 of Treponema pallidum, an agent of syphilis. Immunofluorescence assay and immunogold labeling with anti-Td92 Ab revealed that Td92 had surface-exposed epitopes. And Td92 was capable of binding to fibronectin and KB cells, an oral epithelial cell line. In addition, Td92 could enter the KB cells. These results indicate that Td92 is a fibronectin-binding protein which can bind to and internalize into the host cells, facilitating the virulence of T. denticola.