Immunolocalization of a Drosophila phosphatidylinositol transfer protein (rdgB) in normal and rdgA mutant photoreceptor cells with special reference to the subrhabdomeric cisternae.
AuthorsSuzuki E, Hirosawa K
JournalJ Electron Microsc (Tokyo)
PubMed ID7996077
'Distribution of rdgB protein, which was recently shown to be a Drosophila phosphatidylinositol transfer protein, was studied in the photoreceptor cells of compound eyes of normal and rdgAPC47 mutant of Drosophila melanogaster by immunoelectron microscopy using (1) pre-embedding HRP staining, (2) pre-embedding NANOGOLD labeling followed by siliver enhancement, (3) and ... More
A practical technique to postfix nanogold-immunolabeled specimens with osmium and to embed them in Epon for electron microscopy.
AuthorsSawada H, Esaki M
JournalJ Histochem Cytochem
PubMed ID10727291
'Nanogold is a tiny gold probe, freely diffusible in cells and tissues, and is suitable for pre-embedding immunohistochemistry. However, it is necessary to develop Nanogold to a larger size so that it can be observed by conventional transmission electron microscopy. Silver enhancement is usually used for visualizing Nanogold, but the ... More
Synaptotagmin VII as a plasma membrane Ca(2+) sensor in exocytosis.
AuthorsSugita S, Han W, Butz S, Liu X, Fernández-Chacón R, Lao Y, Südhof TC
JournalNeuron
PubMed ID11395007
'Synaptotagmins I and II are Ca(2+) binding proteins of synaptic vesicles essential for fast Ca(2+)-triggered neurotransmitter release. However, central synapses and neuroendocrine cells lacking these synaptotagmins still exhibit Ca(2+)-evoked exocytosis. We now propose that synaptotagmin VII functions as a plasma membrane Ca(2+) sensor in synaptic exocytosis complementary to vesicular synaptotagmins. ... More
Cryo-electron microscopic localization of protein L7/L12 within the Escherichia coli 70 S ribosome by difference mapping and Nanogold labeling.
AuthorsMontesano-Roditis L, Glitz DG, Traut RR, Stewart PL
JournalJ Biol Chem
PubMed ID11278411
'The Escherichia coli ribosomal protein L7/L12 is central to the translocation step of translation, and it is known to be flexible under some conditions. The assignment of electron density to L7/L12 was not possible in the recent 2.4 A resolution x-ray crystallographic structure (Ban, N., Nissen, P., Hansen, J., Moore, ... More
Highly sensitive DNA, RNA and antigen detection methods: streptavidin-Nanogold-silver staining.
AuthorsHauser-Kronberger C
JournalCell Vis
PubMed ID9660736
Labeling with nanogold and undecagold: techniques and results.
AuthorsHainfeld JF
JournalScanning Microsc Suppl
PubMed ID9601549
A significant new development in gold labeling for microscopy has been achieved through the use of gold cluster compounds that are covalently attached to antibodies or other probe molecules. These unique gold probes are smaller than most colloidal gold conjugates and exhibit improved penetration into tissues, higher labeling densities, and ... More
Use of nanogold followed by silver enhancement and gold toning for preembedding immunolocalization in osmium-fixed, Epon-embedded tissues.
AuthorsSawada H, Esaki M
JournalJ Electron Microsc (Tokyo)
PubMed ID7722428
A reliable, sensitive, high-resolution method with good structural visualization for preembedding immunoelectron microscopy was proposed. The present technique involves the following processes. Immunolabeling of cryostat sections with primary antibodies and with nanogolds, silver intensification for visualization of nanogold secondary antibodies, gold-toning for stabilization of silver shells, and osmium postfixation and ... More
Peculiar distribution of fodrin in fat-storing cells.
AuthorsAoki T, Hagiwara H, Fujimoto T
JournalExp Cell Res
PubMed ID9260899
Fat-storing cells (FSCs) show unique morphology containing many lipid droplets in the cytoplasm. In this study, we found that a membrane skeletal protein, fodrin, shows peculiar distribution in FSCs of rat liver. By immunofluorescence microscopy of FSCs in culture, intense labeling for fodrin was seen as coarse filaments in the ... More
High performance Nanogold-silver in situ hybridisation.
AuthorsHacker GW
JournalEur J Histochem
PubMed ID9728288
Conventional in situ hybridisation (ISH) usually requires the presence of at least 10-50 copies of the nucleic acid sequence in question per cell. In situ PCR has been proposed as an alternative method, which may yield single-copy sensitivity, but shows a relatively high rate of false-negative or even false-positive reactions. ... More
Lysines 128 and 132 enable lipopolysaccharide binding to MD-2, leading to Toll-like receptor-4 aggregation and signal transduction.
AuthorsVisintin A, Latz E, Monks BG, Espevik T, Golenbock DT
JournalJ Biol Chem
PubMed ID12960171
Three cell-surface proteins have been recognized as components of the mammalian signaling receptor for bacterial lipopolysaccharide (LPS): CD14, Toll-like receptor-4 (TLR4), and MD-2. Biochemical and visual studies shown here demonstrate that the role of CD14 in signal transduction is to enhance LPS binding to MD-2, although its expression is not ... More
The use of electron microscopic immunocytochemistry with silver-enhanced 1.4-nm gold particles to localize GAD in the cerebellar nuclei.
AuthorsGilerovitch HG, Bishop GA, King JS, Burry RW
JournalJ Histochem Cytochem
PubMed ID7868863
Silver enhancement of small gold particles can be used with pre-embedding immunocytochemistry to analyze the distribution of label over cell organelles. We have developed a method that improves tissue morphology, has good penetration of reagents, and allows greater control of silver enhancement of 1.4-nm gold. In this study we analyzed ... More
Direct comparison of the sensitivity of enzyme histochemical and immunohistochemical methods: cathepsin B expression in human colorectal mucosa.
AuthorsBleeker FE, Hazen LG, Köhler A, Van Noorden CJ
JournalActa Histochem
PubMed ID10990063
Immunohistochemical localization of the proteinase cathepsin B has been compared directly with localization of cathepsin B activity with a catalytic (enzyme) histochemical method. The 2 approaches demonstrate principally different aspects of an enzyme. The immunohistochemical method localizes the enzyme protein whether it is active or not whereas the catalytic method ... More
Combined fluorescent and gold immunoprobes: reagents and methods for correlative light and electron microscopy.
AuthorsPowell RD, Halsey CM, Hainfeld JF
JournalMicrosc Res Tech
PubMed ID9712158
Immunoprobes which incorporate both a fluorescent label and a 1.4 nm gold cluster compound were prepared by covalent conjugation to Fab' antibody fragments of the Nanogold cluster label followed by a fluorescent moiety. These new immunoconjugates allow the collection of two complementary sets of data, from fluorescence and electron microscopy, ... More
Introduction of a novel HRP substrate-Nanogold probe for signal amplification in immunocytochemistry.
AuthorsMayer G, Leone RD, Hainfeld JF, Bendayan M
JournalJ Histochem Cytochem
PubMed ID10727287
Amplification of immunological signals with catalyzed reporter deposition (CARD) allows improved detection of scarce tissue antigens in light and electron microscopy. The technique takes advantage of the oxidation ability of horseradish peroxidase (HRP), in the presence of hydrogen peroxide, to yield the accumulation of one of its specific reporter-tagged substrates. ... More
Ultrastructural localization of beta-actin and amphoterin mRNA in cultured cells: application of tyramide signal amplification and comparison of detection methods.
AuthorsPunnonen EL, Fages C, Wartiovaara J, Rauvala H
JournalJ Histochem Cytochem
PubMed ID9857217
We describe a nonradioactive preembedding in situ hybridization protocol using digoxigenin-labeled RNA probes and tyramide signal amplification to increase the sensitivity of detection. The protocol is sensitive enough for electron microscopic localization of endogenous messenger RNAs encoding beta-actin and amphoterin. Three visualization methods were compared: diaminobenzidine enhanced by nickel, Nanogold ... More
Lipopolysaccharide internalization activates endotoxin-dependent signal transduction in cardiomyocytes.
AuthorsCowan DB, Noria S, Stamm C, Garcia LM, Poutias DN, del Nido PJ, McGowan FX
JournalCirc Res
PubMed ID11249872
We tested the hypothesis that bacterial lipopolysaccharide (LPS) must be internalized to facilitate endotoxin-dependent signal activation in cardiac myocytes. Fluorescently labeled LPS was used to treat primary cardiomyocyte cultures, perfused heart preparations, and the RAW264.7 macrophage cell line. Using confocal microscopy and spectrofluorometry, we found that LPS was rapidly internalized ... More
Action of gold chloride ("gold toning") on silver-enhanced 1 nm gold markers.
AuthorsPohl K, Stierhof YD
JournalMicrosc Res Tech
PubMed ID9712163
In conventional immunoelectron microscopy (IEM), very small colloidal gold particles (0.8-3 nm), or the gold compound Nanogold (1.4 nm) are silver-enhanced for easy detection. However, silver enhancement has drawbacks. First, the silver layer is dissolved during fixation with osmium tetroxide, even if the concentration and incubation time are strongly reduced ... More
Detection of human papillomavirus in cervical carcinoma: comparison of peroxidase, Nanogold, and catalyzed reporter deposition (CARD)-Nanogold in situ hybridization.
We compared three in situ hybridization (ISH) methods for their applicability and sensitivity in detecting human papillomavirus (HPV) in 61 cases (1 Grade 1, 18 Grade 2, 42 Grade 3) of routinely processed squamous cell cervical carcinoma. A commercially available biotinylated probe for HPV-16/18 was applied to serial sections and ... More
Immunocytochemistry of extracellular matrix components in the rat seminiferous tubule: electron microscopic localization with improved methodology.
AuthorsYazama F, Esaki M, Sawada H
JournalAnat Rec
PubMed ID9143667
BACKGROUND: Cell-cell interactions between Sertoli, myoid, Leydig, and germ cells are thought to be essential for spermatogenesis. These cells interact with each other through the extracellular matrices (ECMs) of the testicular lamina propria, which thus may have an important function in spermatogenesis. For an understanding of the role of ECMs ... More
Signal amplification in immunohistochemistry at the light microscopic level using biotinylated tyramide and nanogold-silver staining.
AuthorsKöhler A, Lauritzen B, Van Noorden CJ
JournalJ Histochem Cytochem
PubMed ID10858270
Signal amplification techniques greatly enhance the sensitivity of immunohistochemical (IHC) and in situ hybridization (ISH) methods. In particular, catalyzed signal amplification (CSA) using labeled tyramide or Nanogold-silver staining is an important signal amplification tool. We have applied a combination of both techniques, as has been introduced for ISH, for a ... More
Clinical relevance of HPV 16/18 testing methods in cervical squamous cell carcinoma.
AuthorsGraf AH, Cheung AL, Hauser-Kornberger C, Dandachi N, Tubbs RR, Dietze O, Hacker GW
JournalAppl Immunohistochem Mol Morphol
PubMed ID11127922
Three different in situ hybridization (ISH) methods were compared for their clinical relevance and suitability in detecting human papillomavirus (HPV) 16/18 in 55 cases of squamous cell carcinoma (SCC) of the uterine cervix. After the initial biopsy, surgery, and/or radiation therapy, patients were followed for 5 to 8 years. A ... More
Silver enhancement of gold antibody probes in pre-embedding electron microscopic immunocytochemistry.
AuthorsBurry RW, Vandré DD, Hayes DM
JournalJ Histochem Cytochem
PubMed ID1453003
In pre-embedding EM immunocytochemistry with gold probes, the gold must be small enough to penetrate through cell membranes treated with mild detergents. Antibodies labeled with small gold probes (1-1.4 nm) are too small to be resolved in thin sections but can be seen if they are silver-enhanced after the gold ... More
Fixation conditions for DNA and RNA in situ hybridization: a reassessment of molecular morphology dogma.
Neutral buffered formalin (NBF) (4% neutral buffered formaldehyde) has been advocated by most investigators as the primary fixative of choice for in situ hybridization (ISH), and specific anecdotal cautions interdicting the use of precipitating fixatives, which otherwise may offer certain advantages such as superior nuclear detail, are common. Few systematic ... More