Sensitive in situ hybridization with catalyzed reporter deposition, streptavidin-Nanogold, and silver acetate autometallography: detection of single-copy human papillomavirus.
AuthorsZehbe I, Hacker GW, Su H, Hauser-Kronberger C, Hainfeld JF, Tubbs R
JournalAm J Pathol
PubMed ID9137082
The usefulness of standard in situ hybridization for viral nucleic acid detection is occasionally limited by its sensitivity limit of 10 to 50 copies per cell. A modified version of the recently described signal amplification method, catalyzed reporter deposition (CARD), and its application to formalin-fixed cells and tissue sections is ... More
In situ polymerase chain reaction amplification of HIV-1 DNA in brain tissue.
AuthorsStrappe PM, Wang TH, McKenzie CA, Lowrie S, Simmonds P, Bell JE
JournalJ Virol Methods
PubMed ID9562406
A direct in situ polymerase chain reaction (IS-PCR) assay is described for the detection of HIV-1 proviral DNA in formalin fixed paraffin embedded brain tissue. Biotin-16-dUTP is incorporated during the PCR process and microwave pretreatment of tissue sections ensures that no non-specific incorporation into damaged or nicked genomic DNA occurs. ... More
Attenuation of astroglial reactivity by interleukin-10.
AuthorsBalasingam V, Yong VW
JournalJ Neurosci
PubMed ID8622125
'Prominent responses that follow brain trauma include the activation of microglia, recruitment of blood-derived macrophages, and astroglial reactivity. Based on evidence that cytokines produced by macrophages/microglia may cause astrocytes to become reactive, the aim of this study was to determine whether astroglial reactivity could be attenuated by interleukin (IL)-10, a ... More
Biotinyl-tyramide: a novel approach for electron microscopic immunocytochemistry.
AuthorsMayer G, Bendayan M
JournalJ Histochem Cytochem
PubMed ID9358846
'The biotinyl-tyramide protocol recently introduced for sensitive light microscopic immunocytochemistry was applied to electron microscopy and revealed various tissue antigens with high resolution. The protocol consists of an indirect method in which thin tissue sections are incubated successively within a specific primary antibody, followed by a biotinylated secondary antibody, streptavidin-HRP, ... More
Galanin-5-hydroxytryptamine interactions: electrophysiological, immunohistochemical and in situ hybridization studies on rat dorsal raphe neurons with a note on galanin R1 and R2 receptors.
AuthorsXu ZQ, Zhang X, Pieribone VA, Grillner S, Hökfelt T
JournalNeuroscience
PubMed ID9722143
'Galaninergic mechanisms related to 5-hydroxytryptamine neurons in the dorsal raphe nucleus of the rat were analysed using electrophysiology, immunohistochemistry and in situ hybridization. Galanin caused a dose-dependent hyperpolarization accompanied by a decrease in membrane resistance in most 5-hydroxytryptamine-sensitive dorsal raphe neurons. The galanin-induced outward current reversed at about - 105 ... More
Co-localization of the glial cell-line derived neurotrophic factor and its functional receptor c-RET in a subpopulation of rat dorsal root ganglion neurons.
AuthorsHonda T, Takahashi M, Sugiura Y
JournalNeurosci Lett
PubMed ID10554981
'Co-localization of the glial cell line-derived neurotrophic factor (GDNF) and its functional receptor c-RET was examined immunohistochemically in the dorsal root ganglion (DRG) of the 5th lumbar nerve in rats. The total 1728 DRG neurons were observed in three female rats (Wistar) with ca. 200 g body weight, 46.6% of ... More
Catalyzed reporter deposition, a novel method of signal amplification. II. Application to membrane immunoassays.
AuthorsBobrow MN, Shaughnessy KJ, Litt GJ
JournalJ Immunol Methods
PubMed ID1849153
'In a previous publication (Bobrow et al., J. Immunol. Methods (1989) 279-285), we described a novel signal amplification method, catalyzed reporter deposition (CARD), and its application to microplate immunoassays. The method utilizes the analyte-dependent reporter enzyme (ADRE) to catalyze the deposition of additional reporter onto the surface of a solid-phase ... More
Cryopreservation of organotypic multicellular spheroids from human gliomas.
AuthorsKaaijk P, van den Berg F, Van Amstel P, Troost D
JournalNeuropathol Appl Neurobiol
PubMed ID9004246
'Fresh human glioma tissue can be cultured on agarose to form organotypic multicellular spheroids (OMS). The major advantage of OMS is the preservation of the cellular heterogeneity and the tumour architecture, which is lost in conventional monolayer cultures. The present study was undertaken to assess the possibilities of storing frozen ... More
Sensitive multicolor fluorescence in situ hybridization using catalyzed reporter deposition (CARD) amplification.
AuthorsSpeel EJ, Ramaekers FC, Hopman AH
JournalJ Histochem Cytochem
PubMed ID9313806
'We describe the simultaneous localization of DNA sequences in cell and chromosome preparations by means of differently fluorochrome-labeled (AMCA, FITC, TRITC) tyramides using the catalyzed reporter deposition (CARD) procedure. For this purpose, repeated as well as single-copy DNA probes were labeled with biotin, digoxigenin, and FITC, hybridized, and visualized with ... More
'To gain a molecular understanding of tumor angiogenesis, we compared gene expression patterns of endothelial cells derived from blood vessels of normal and malignant colorectal tissues. Of over 170 transcripts predominantly expressed in the endothelium, 79 were differentially expressed, including 46 that were specifically elevated in tumor-associated endothelium. Several of ... More
Enhancement of immunohistochemical detection of HIV-1 p24 antigen in brain by tyramide signal amplification.
AuthorsStrappe PM, Wang TH, McKenzie CA, Lowrie S, Simmonds P, Bell JE
JournalJ Virol Methods
PubMed ID9274823
'Human immunodeficiency virus type 1 (HIV-1) infection of the brain has been demonstrated in formalin fixed, paraffin embedded post-mortem brain tissue (PM) by chromogenic immunohistochemistry for the HIV p24 antigen. The sensitivity of antigen detection is increased significantly by tyramide signal amplification (TSA) compared to the conventional peroxidase labelled Avidin-Biotin ... More
Studies on the phenotype of migrant thymic stem cells.
AuthorsSuniara RK, Jenkinson EJ, Owen JJ
JournalEur J Immunol
PubMed ID9933088
'Stem cells first enter the thymus around the 11th to 12th days of gestation in BALB/c mouse embryos. The phenotype of these stem cells has been difficult to determine because their entry occurs when the thymic primordium is very small and involves too few stem cells to allow studies by ... More
Post-incubation heating significantly improves tyramide signal amplification.
AuthorsVolante M, Pecchioni C, Bussolati G
JournalJ Histochem Cytochem
PubMed ID11185079
A highly sensitive nonisotopic detection method for in situ hybridization.
AuthorsHolm R
JournalAppl Immunohistochem Mol Morphol
PubMed ID10937065
'In situ hybridization is a technique that allows detection of specific DNA and RNA sequences in tissue sections. Nonisotopic techniques are fast and give a precise localization of the hybridization product, but a drawback is the low sensitivity. However, the sensitivity is dependent on the detection system used. To evaluate ... More
Human immunodeficiency virus type 1 p24 concentration measured by boosted ELISA of heat-denatured plasma correlates with decline in CD4 cells, progression to AIDS, and survival: comparison with viral RNA measurement.
'Human immunodeficiency virus type 1 (HIV-1) RNA and p24 antigen concentrations were determined in plasma samples from 169 chronically infected patients (median CD4 cell count, 140 cells/microL; range, 0-1500 cells/microL). p24 quantification involved heat-mediated immune complex dissociation and tyramide signal amplification-boosted ELISA, which has a diagnostic sensitivity similar to that ... More
Use of methacrylate De-embedding protocols for in situ hybridization on semithin plastic sections with multiple detection strategies.
AuthorsWarren KC, Coyne KJ, Waite JH, Cary SC
JournalJ Histochem Cytochem
PubMed ID9446821
'New plastic resins are gradually replacing traditional paraffin-embedding in situ hybridization (ISH) strategies. One unique resin that has not been fully investigated or exploited with respect to light microscopic ISH is a methacrylate mixture. The advantage of this resin is its ability to be removed from tissues postsectioning, dramatically increasing ... More
The visual assignment of genes by fiber-fish: BTF3 protein homologue gene (BTF3) and a novel pseudogene of human RNA helicase A (DDX9P) on 13q22.
AuthorsKlockars T, Isosomppi J, Laan M, Kakko N, Palotie A, Peltonen L
JournalGenomics
PubMed ID9325059
'Chromosome 13 is one of the poorly mapped human chromosomes. As an example, only two cloned genes have been assigned to bands 13q22-q31. Our characterization of the critical region for the variant form of late infantile neuronal ceroid lipofuscinosis (vLINCL, locus definition CLN5) disease region on 13q22 resulted in the ... More
Expression of galanin and a galanin receptor in several sensory systems and bone anlage of rat embryos.
AuthorsXu ZQ, Shi TJ, Hökfelt T
JournalProc Natl Acad Sci U S A
PubMed ID8962153
'Using in situ hybridization and immunohistochemistry the expression of, respectively, prepro-galanin (pre-pro-GAL) mRNA and GAL receptor-1 mRNA, as well as GAL-like and GAL message-associated peptide-like immunoreactivities, were studied in rats from embryonic day 14 (E14) to postnatal day 1. GAL expression was observed already at E14 in trigeminal and dorsal ... More
Immunocytochemistry of endothelial nitric oxide synthase in the rat brain: a light and electron microscopical study using the tyramide signal amplification technique.
AuthorsStanarius A, Töpel I, Schulz S, Noack H, Wolf G
JournalActa Histochem
PubMed ID9429601
'There are many inconsistencies in the literature about the cellular and subcellular distribution of the endothelial isoform of nitric oxide synthase (eNOS) in the brain. We have re-investigated its localization by light and electron microscopical (LM, EM) immunocytochemistry and the NADPH-diaphorase reaction. Using bovine aortic tissue as a positive control ... More
Cocaine- and amphetamine-regulated transcript in the rat vagus nerve: A putative mediator of cholecystokinin-induced satiety.
AuthorsBroberger C, Holmberg K, Kuhar MJ, Hökfelt T
JournalProc Natl Acad Sci U S A
PubMed ID10557351
'Cocaine- and amphetamine-regulated transcript (CART) is widely expressed in the central nervous system. Recent studies have pointed to a role for CART-derived peptides in inhibiting feeding behavior. Although these actions have generally been attributed to hypothalamic CART, it remains to be determined whether additional CART pathways exist that link signals ... More
The detection of CD2+, CD4+, CD8+, and WC1+ T lymphocytes, B cells and macrophages in fixed and paraffin embedded bovine tissue using a range of antigen recovery and signal amplification techniques.
AuthorsGutierrez M, Forster FI, McConnell SA, Cassidy JP, Pollock JM, Bryson DG
JournalVet Immunol Immunopathol
PubMed ID10587310
'In order to develop procedures to label the main bovine leucocyte populations in paraffin embedded sections, the immunoreactivity of 25 monoclonal antibodies (mAbs) to different leucocyte antigens was assessed with formal dichromate (FD5) and 10% formalin fixation, a battery of antigen retrieval (AR) methods, and the biotin-tyramide amplification system. All ... More
Nonsaturable binding indicates clustering of tau on the microtubule surface in a paired helical filament-like conformation.
AuthorsAckmann M, Wiech H, Mandelkow E
JournalJ Biol Chem
PubMed ID10869348
'Tau protein modulates microtubule dynamics and forms insoluble aggregates in Alzheimer''s disease. Because there is a discrepancy between reported affinities of Tau to microtubules, we determined the interaction over a wide concentration range using a sensitive enzyme-linked immunosorbent assay. We found that the interaction is biphasic and not monophasic as ... More
Subtypes Y1 and Y2 of the neuropeptide Y receptor are respectively expressed in pro-opiomelanocortin- and neuropeptide-Y-containing neurons of the rat hypothalamic arcuate nucleus.
AuthorsBroberger C, Landry M, Wong H, Walsh JN, Hökfelt T
JournalNeuroendocrinology
PubMed ID9430445
'The arcuate nucleus of the hypothalamus houses a number of neurochemically different cell populations. Among these, a dense cluster of small neuropeptide-Y (NPY)-expressing neurons is located in its ventromedial subdivision and a pro-opiomelanocortin (POMC)-expressing neuron population in its ventrolateral part. Furthermore, both neuropeptide Y Y1 and Y2 receptors (Y1-Rs and ... More
Vasodepressor effects of exercise are accompanied by reduced circulating ouabainlike immunoreactivity and normalization of nitric oxide synthesis.
AuthorsKomiyama Y, Kimura Y, Nishimura N, Hara K, Mori T, Okuda K, Munakata M, Masuda M, Murakami T, Takahashi H
JournalClin Exp Hypertens
PubMed ID9107442
'Our object was to evaluate the effects of regular mild exercise on blood pressure and on circulating level of ouabainlike factors (OLF) and of nitrate anion, an endproduct of nitric oxide (NO) in humans. We measured plasma ouabainlike immunoreactivity (OLI) and nitrate ions (NO3.) before and after mild exercise for ... More
Simultaneous detection of two independent antigens by double staining with two mouse monoclonal antibodies.
AuthorsTeramoto N, Szekely L, Pokrovskaja K, Hu LF, Yoshino T, Akagi T, Klein G
JournalJ Virol Methods
PubMed ID9705180
'Simultaneous detection of two antigens by immunostaining usually requires primary antibodies from two different species or a hapten modification of one of the antibodies if they are from the same species. A novel double staining method is described for immunodetection of two independent antigens using two mouse monoclonal antibodies. The ... More
The ubiquitin conjugation system is required for ligand-induced endocytosis and degradation of the growth hormone receptor.
AuthorsStrous GJ, van Kerkhof P, Govers R, Ciechanover A, Schwartz AL
JournalEMBO J
PubMed ID8670885
'The ubiquitin-dependent protein degradation system has recently been implicated in downregulation of signal transducing receptors. Growth hormone receptor (GHR) cDNA was transfected into Chinese hamster ovary cells, which exhibit a temperature-sensitive defect in ubiquitin conjugation (CHO-ts20), as well as into wild-type cells (CHO-E36). Upon binding of growth hormone (GH), two ... More
Tyramide signal amplification (TSA)-FISH applied to mapping PCR-labeled probes less than 1 kb in size.
AuthorsSchriml LM, Padilla-Nash HM, Coleman A, Moen P, Nash WG, Menninger J, Jones G, Ried T, Dean M
JournalBiotechniques
PubMed ID10489619
'Tyramide signal amplification (TSA)-FISH was used to map one mouse and two human DNA probes of less than 1 kb in size. The two human probes were 319 and 608 bp, and the mouse probe was 855 bp. Probes, made from PCR products, were labeled by incorporating biotin-11-dUTP (human) and ... More
Catalyzed reporter deposition, a novel method of signal amplification. Application to immunoassays.
AuthorsBobrow MN, Harris TD, Shaughnessy KJ, Litt GJ
JournalJ Immunol Methods
PubMed ID2558138
'A novel signal amplification method, catalyzed reporter deposition (CARD), and its application to immunoassays is described. The method involves utilizing an analyte-dependent reporter enzyme (ADRE) to catalyze the deposition of additional reporter on the surface in a solid-phase immunoassay. In the examples described, deposition of reporter is facilitated by using ... More
Synthesis and purification of horseradish peroxidase-labeled oligonucleotides for tyramide-based fluorescence in situ hybridization.
Authorsvan Gijlswijk RP, van de Corput MP, Bezrookove V, Wiegant J, Tanke HJ, Raap AK
JournalHistochem Cell Biol
PubMed ID10817671
'A method is presented to conjugate horseradish peroxidase (HRP) to oligodeoxynucleotides for fluorescence in situ hybridization assays employing tyramide signal amplification (TSA). HRP is covalently bound to the oligonucleotide by thiol ether linkage and purified by high-performance liquid chromatography. With TSA detection, a single HRP-labeled oligonucleotide probe is sufficient for ... More
Transformation of primary human endothelial cells by Kaposi's sarcoma-associated herpesvirus.
AuthorsFlore O, Rafii S, Ely S, O'Leary JJ, Hyjek EM, Cesarman E
JournalNature
PubMed ID9707121
'Kaposi''s sarcoma-associated herpesvirus (KSHV), or human herpesvirus 8, is invariably present in Kaposi''s sarcoma lesions. KSHV contains several viral oncogenes and serological evidence suggests that KSHV infection is necessary for the development of Kaposi''s sarcoma, but cellular transformation by this virus has not so far been demonstrated. KSHV is found ... More
cFos immunoreactivity is enhanced with biotin amplification.
AuthorsBerghorn KA, Bonnett JH, Hoffman GE
JournalJ Histochem Cytochem
PubMed ID7983364
'Through modification of the protocol by Adams, we developed a biotin amplification procedure for immunofluorescence staining of immediate early gene proteins and also applied biotin amplification for metal enhancement of diaminobenzidine staining in an immunoperoxidase protocol. Commercially available anti-cFos antisera were used to compare conventional "Elite" avidin-biotin complex reactions with ... More
Dual fluorescent in situ hybridization and immunohistochemical detection with tyramide signal amplification.
AuthorsZaidi AU, Enomoto H, Milbrandt J, Roth KA
JournalJ Histochem Cytochem
PubMed ID10990490
'To understand the biological relationships among various molecules, it is necessary to define the cellular expression patterns of multiple genes and gene products. Relatively simple methods for performing multi-label immunohistochemical detection are available. However, there is a paucity of techniques for dual immunohistochemical (IHC) and mRNA in situ hybridization (ISH) ... More
Ultra-sensitive FISH is a useful tool for studying chronic HIV-1 infection.
AuthorsDeichmann M, Bentz M, Haas R
JournalJ Virol Methods
PubMed ID9128858
'Using an ultra-sensitive fluorescence in situ hybridization (FISH) protocol, integration sites of the human immunodeficiency virus-1 (HIV-1) were mapped in two model cell lines for chronic HIV infection. In the promonocytic cell line U1/HIV-1, the two integrated copies of proviral HIV-DNA were located on the short arm of one copy ... More
Assignment of human MFNG, manic fringe Drosophila homolog, to 22q13.1 using tyramide fluorescence in situ hybridization (T-FISH).
AuthorsVan Tine BA, Knops J, Shaw GM, May WA
JournalCytogenet Cell Genet
PubMed ID10640833
9 enhanced tyramide signal amplification immunohistochemical detection
AuthorsRoth KA, Adler K, Bobrow MN,
JournalJ Histochem Cytochem
PubMed ID10567456
Tech.Sight. Worth its weight in gold.
AuthorsBendayan M
JournalScience
PubMed ID11233453
Tyramide amplification allows anterograde tracing by horseradish peroxidase-conjugated lectins in conjunction with simultaneous immunohistochemistry
AuthorsKressel M
JournalJ Histochem Cytochem
PubMed ID9524199
LacZ staining in paraffin-embedded tissue sections.
AuthorsHendrikx PJ, Vermeulen J, Hagenbeek A, Vermey M, Martens AC
JournalJ Histochem Cytochem
PubMed ID8918907
Femora and tibiae of rats carrying leukemia from a LacZ-marked acute promyelocytic leukemia-derived leukemic cell line (LT12NL15) were decalcified using EDTA and routinely embedded in paraffin. Sections were used to develop for the first time an immunostaining method for LacZ, employing catalyzed reporter deposition (CARD) based on the deposition of ... More
Dehydration modifies somal CRH immunoreactivity in the rat hypothalamus: an immunocytochemical study in the absence of colchicine.
AuthorsKay-Nishiyama C, Watts AG
JournalBrain Res
PubMed ID10082905
Corticotropin-releasing hormone immunoreactive (CRH-ir) neurons were examined in the hypothalamus of euhydrated and dehydrated rats without using colchicine. CRH-ir cells were observed in the lateral hypothalamus, retrochiasmatic, and in magnocellular parts of the supraoptic and paraventricular hypothalamic nucleus (PVH) in dehydrated but not euhydrated animals. However, CRH-ir neurons were decreased ... More
Fluorochrome-labeled tyramides: use in immunocytochemistry and fluorescence in situ hybridization.
The peroxidase-mediated deposition of hapten- and fluorochrome-labeled tyramides has recently been shown to increase the sensitivity of immunofluorescence and fluorescence in situ hybridization techniques. We have evaluated a number of red, green, and blue fluorescent tyramides for detection of antigens in tissue sections and cytospin preparations and for the detection ... More
Tyramide signal amplification in brain immunocytochemistry: adaptation to electron microscopy.
AuthorsStanarius A, Faber-Zuschratter H, Töpel I, Schulz S, Wolf G
JournalJ Neurosci Methods
PubMed ID10379579
The tyramide signal amplification (TSA) technique is well-established in light microscopic immunohistochemistry and in situ hybridization to improve the signal-to-noise ratio. The present study deals with its adaptation to the electron microscopic level using the pre-embedding technique and a modified protocol. The outcome of immunolabeling of most of the antigens ... More
Improved sensitivity of whole-cell hybridization by the combination of horseradish peroxidase-labeled oligonucleotides and tyramide signal amplification.
AuthorsSchönhuber W, Fuchs B, Juretschko S, Amann R
JournalAppl Environ Microbiol
PubMed ID9251215
The substrate fluorescein-tyramide was combined with oligonucleotide probes directly labeled with horseradish peroxidase to improve the sensitivity of in situ hybridization of whole fixed bacterial cells. Flow cytometry and quantitative microscopy of cells hybridized by this technique showed 10- to 20-fold signal amplifications relative to fluorescein-monolabeled probes. The application of ... More
Detection of apoptotic DNA damage in prostate hyperplasia using tyramide-amplified avidin-HRP.
AuthorsSlater M, Murphy CR
JournalHistochem J
PubMed ID10646840
Avidin binds to damaged DNA with high specificity. Avidin conjugated to horseradish peroxidase may therefore be used to label apoptotic DNA damage, using standard immunohistochemical protocols. However, the resulting label may be too weak to visualise. We used tyramide signal amplification to enhance the avidin-peroxidase signal in a rat model ... More
An in situ hybridization protocol to detect rare mRNA expressed in neural tissue using biotin-labelled oligonucleotide probes.
AuthorsTata AM
JournalBrain Res Brain Res Protoc
PubMed ID11223418
The use of the non-radioactive in situ hybridization protocols has allowed in general to obtain a better resolution of different transcripts at histological and cytological levels with a shortening of the developmental time. The common protocols using digoxigenin and biotin-labelled probes share a considerable limitation depending on the amount of ... More
Amplification methods to increase the sensitivity of in situ hybridization: play card(s).
AuthorsSpeel EJ, Hopman AH, Komminoth P
JournalJ Histochem Cytochem
PubMed ID10026231
In situ hybridization (ISH) has proved to be an invaluable molecular tool in research and diagnosis to visualize nucleic acids in their cellular environment. However, its applicability can be limited by its restricted detection sensitivity. During the past 10 years, several strategies have been developed to improve the threshold levels ... More
Specific in situ hybridization of Haemobartonella felis with a DNA probe and tyramide signal amplification.
AuthorsBerent LM, Messick JB, Cooper SK, Cusick PK
JournalVet Pathol
PubMed ID10643980
Haemobartonella felis is an epierythrocytic bacterium suspected to be the causative agent of feline infectious anemia. Previous studies with a polymerase chain reaction assay have identified a mycoplasmal 16S rRNA gene sequence that coincides with clinical disease and the presence of organisms in the blood. Tissues from a cat experimentally ... More
Expression of CaT-like, a novel calcium-selective channel, correlates with the malignancy of prostate cancer.
AuthorsWissenbach U, Niemeyer BA, Fixemer T, Schneidewind A, Trost C, Cavalie A, Reus K, Meese E, Bonkhoff H, Flockerzi V
JournalJ Biol Chem
PubMed ID11278579
The regulation of intracellular Ca(2+) plays a key role in the development and growth of cells. Here we report the cloning and functional expression of a highly calcium-selective channel localized on the human chromosome 7. The sequence of the new channel is structurally related to the gene product of the ... More
Neuronal nitric oxide synthase in the vasculature of the rat brain: an immunocytochemical study using the tyramide signal amplification technique.
AuthorsStanarius A, Seidel B, Wolf G
JournalJ Neurocytol
PubMed ID10640188
Using the highly sensitive tyramide-signal-amplification technique, we examined immunocytochemically the distribution of neuronal nitric oxide synthase (nNOS) in blood vessels of the rat brain. In contrast to the endothelial isoform, no clear-cut immunostaining could be obtained for the nNOS at the light microscopic level. An occasional faint immunoreaction at the ... More
Improved localization of fluorescent tyramides for fluorescence in situ hybridization using dextran sulfate and polyvinyl alcohol.
Recently, a peroxidase-mediated amplification system has been described for immunofluorescence and fluorescence in situ hybridization studies. It is based on the deposition of hapten- or fluorochrome-labeled tyramide molecules. Although providing a significantly increased detection sensitivity compared to conventional procedures, its localization properties are inferior because of free diffusion of intermediate ... More
Focal sites of DNA repair synthesis in human chromosomes.
Nucleotide excision repair (NER) is the principle pathway by which the human cells eliminate UV-induced lesions from their genomic DNA. The process can be visualized through the labelling of the nucleotides that are incoporated into repair patches, following the excision of the damaged stretch of DNA. In this study we ... More
High-resolution physical and transcriptional mapping of the autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy locus on chromosome 21q22.3 by FISH.
AuthorsAaltonen J, Horelli-Kuitunen N, Fan JB, Björses P, Perheentupa J, Myers R, Palotie A, Peltonen L
JournalGenome Res
PubMed ID9267805
Autoimmune-polyendocrinopathy-candidiasis-ectodermal dystrophy (APECED, PGD type I) is an autosomal recessive disease enriched in the Finnish population. Previously, we have assigned APECED to a 2.6-cM interval on chromosome 21q22.3 by linkage analysis in 14 Finnish families. This subtelomeric region of 21q22.3 seems to have sequence features resulting in its under-representation in ... More
High sensitivity detection of HPV-16 in SiHa and CaSki cells utilizing FISH enhanced by TSA.
AuthorsAdler K, Erickson T, Bobrow M
JournalHistochem Cell Biol
PubMed ID9387924
Detection of integrated human papillomavirus type 16 (HPV-16) DNA in SiHa and CaSki cells was used as a model system to demonstrate sensitivity and resolution of a well defined target. Using 293- to 1987-base polymerase chain reaction (PCR)-synthesized probes to the E6 and E7 open reading frames of HPV-16, several ... More
Distribution of the endothelial constitutive nitric oxide synthase in the developing rat brain: an immunohistochemical study.
AuthorsTöpel I, Stanarius A, Wolf G
JournalBrain Res
PubMed ID9554947
The present study deals with the distribution of endothelial constitutive nitric oxide synthase (ecNOS) in the developing rat brain using optimized protocols for preparation and fixation and the tyramide-signal-amplification technique. The immunostaining patterns of a monoclonal antibody against ecNOS are compared with results obtained with a rat pan-endothelial marker, the ... More
Detection of herpes simplex virus DNA in maternal breast milk by in situ hybridization with tyramide signal amplification.
AuthorsKotronias D, Kapranos N
JournalIn Vivo
PubMed ID10757038
In this study we investigated the prevalence of herpes simplex virus 1 (HSV-1) and 2 (HSV-2) in maternal breast milk using the technique of in situ hybridization combined with the sensitive detection system of tyramide signal amplification. Breast milk samples were collected from both breasts of 34 puerperals 4-5 days ... More
In situ hybridization detection of single-copy human papillomavirus on isolated cells, using a catalyzed signal amplification system: GenPoint.
AuthorsLizard G, Démares-Poulet MJ, Roignot P, Gambert P
JournalDiagn Cytopathol
PubMed ID11169889
The performance and drawbacks of GenPoint, which is a catalyzed signal amplification system for immunohistochemistry, have been evaluated for its ability to reveal human papillomavirus (HPV) DNA detected by in situ hybridization with biotinylated DNA probes. For this aim, formalin-fixed cell deposits from carcinoma cells of the uterine cervix, CaSki, ... More
Immunohistochemical detection of the Epstein-Barr virus-encoded latent membrane protein 2A in Hodgkin's disease and infectious mononucleosis.
AuthorsNiedobitek G, Kremmer E, Herbst H, Whitehead L, Dawson CW, Niedobitek E, von Ostau C, Rooney N, Grässer FA, Young LS
JournalBlood
PubMed ID9269787
We describe two new monoclonal antibodies specific for the Epstein-Barr virus (EBV)-encoded latent membrane protein 2A (LMP2A) that are suitable for the immunohistochemical analysis of routinely processed paraffin sections. These antibodies were applied to the immunohistochemical detection of LMP2A in Hodgkin's disease (HD). LMP2A-specific membrane staining was seen in the ... More
Tyramine amplification technique in routine immunohistochemistry.
Authorsvon Wasielewski R, Mengel M, Gignac S, Wilkens L, Werner M, Georgii A
JournalJ Histochem Cytochem
PubMed ID9358847
Signal amplification in immunohistochemistry via binding of biotinylated tyramine to proteins near the site of peroxidase-labeled antibodies is a promising new technique, but studies investigating a wide range of markers are lacking. The tyramine amplification technique (TAT) was investigated on 85 antibodies using a simple and fast protocol, and TAT ... More
Immunogold signal amplification: Application of the CARD approach to electron microscopy.
AuthorsMayer G, Bendayan M
JournalJ Histochem Cytochem
PubMed ID10082744
Catalyzed reporter deposition (CARD) is a technique that allows amplification of routine immunolabeling in light microscopy. This procedure takes advantage of the horseradish peroxidase (HRP) from an HRP-avidin complex to catalyze the accumulation of reporter-conjugated tyramine (a phenolic compound) onto a surface displaying biotinylated antigen-antibody complexes. The large amount of ... More
Co-localization of two different viral genomes in the same sample by double-chemiluminescence in situ hybridization.
AuthorsGentilomi G, Musiani M, Roda A, Pasini P, Zerbini M, Gallinella G, Baraldini M, Venturoli S, Manaresi E,
JournalBiotechniques
PubMed ID9421639
A double-chemiluminescence in situ hybridization has been developed that combines the advantages of chemiluminescence with the detection of two different viral DNAs, i.e., herpes simplex virus (HSV) DNA and cytomegalovirus (CMV) DNA, in infected cells in the same specimen. For the simultaneous detection of these two different viral DNAs, we ... More
In situ hybridization with biotinylated tyramide amplification: detection of human papillomavirus DNA in cervical neoplastic lesions.
AuthorsSano T, Hikino T, Niwa Y, Kashiwabara K, Oyama T, Fukuda T, Nakajima T
JournalMod Pathol
PubMed ID9556418
A novel peroxidase-mediated amplification system, which is based on the deposition of biotinylated tyramide (BT) molecules, was recently described for immunohistochemistry and in situ hybridization (ISH). We report here a highly sensitive ISH system combined with a microwave prehybridization treatment and BT enhancement after hybridization. Our ISH with BT amplification ... More
Automated immunohistochemical staining of formalin-fixed and paraffin-embedded tissues using a catalyzed signal amplification method.
AuthorsHashizume K, Hatanaka Y, Kamihara Y, Tani Y
JournalAppl Immunohistochem Mol Morphol
PubMed ID11277416
An immunohistochemical assay using catalyzed signal amplification (CSA), which is based on the peroxidase catalyzed deposition of biotinylated tyramide, is a highly sensitive method to visualize weak immunohistochemical signals originating from rare antigens or masked antigens in formalin-fixed, paraffin-embedded (FFPE) tissues. However, CSA methods are hampered by poor reproducibility and ... More
Improved mRNA in situ hybridization on formaldehyde-fixed and paraffin-embedded tissue using signal amplification with different haptenized tyramides.
AuthorsSpeel EJ, Saremaslani P, Roth J, Hopman AH, Komminoth P
JournalHistochem Cell Biol
PubMed ID9860255
We report an optimized in situ hybridization (ISH) protocol with a rapid signal amplification procedure based on catalyzed reporter deposition (CARD) to increase the sensitivity of non-isotopic mRNA ISH on formaldehyde-fixed and paraffin-embedded tissue. The CARD method is based on the deposition of haptenized tyramide molecules in the vicinity of ... More
Amplification of fluorescent in situ hybridisation signals in formalin fixed paraffin wax embedded sections of colon tumour using biotinylated tyramide.
AuthorsMcKay JA, Murray GI, Keith WN, McLeod HL
JournalMol Pathol
PubMed ID9536283
Fluorescent in situ hybridisation (FISH) is a powerful tool for the evaluation of chromosomal alterations in formalin fixed paraffin wax embedded sections of colorectal cancer. However, initial experiments using a two-step detection system for digoxigenin labelled chromosome specific centromeric probes resulted in a complete lack of hybridisation signal from a ... More
An enhanced immunocytochemical method for staining bone marrow trephine sections.
AuthorsErber WN, Willis JI, Hoffman GJ
JournalJ Clin Pathol
PubMed ID9215121
AIMS: The detection of cellular antigens in fixed decalcified bone marrow trephine (BMT) sections depends on the method of processing, the nature of the antigen and antibody, antigen retrieval techniques, and the sensitivity of the immunocytochemical method. This study evaluated a tyramide enhanced avidin-biotin immunostaining method on formalin fixed decalcified ... More
Enzyme-based antigen localization and quantitation in cell and tissue samples (Midwestern assay).
AuthorsRoth KA, Brenner JW, Selznick LA, Gokden M, Lorenz RG
JournalJ Histochem Cytochem
PubMed ID9389766
Quantitation of antigen concentration in cell and tissue samples typically requires antigen extraction, which precludes antigen localization in the same sample. Similarly, antigen immunolocalization in fixed cells or tissue sections provides limited information about antigen concentration. We have developed a rapid and sensitive assay for simultaneous antigen localization and quantitation ... More
Peptide-specific T cell clonal expansion in vivo following immunization in the eye, an immune-privileged site.
To visualize the primary antigen-specific T cell response to Ag introduced into the eye, we have used an adoptive transfer system in which a limiting number of OVA peptide (323-339)-specific T cells from a TCR-transgenic mouse were transferred into nonirradiated, syngeneic recipients and then tracked in vivo by staining for ... 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
Visualization of focal nuclear sites of DNA repair synthesis induced by bleomycin in human cells.
In this study, we examined DNA repair synthesis in human cells treated with the radiomimetic drug bleomycin, which efficiently induces double-strand breaks (DSBs). Using tyramide-biotin to amplify fluorescent signals, discrete nuclear foci from the incorporation of 5-iododeoxyuridine (IdU) were detected in proliferating human cells treated with bleomycin. We believe this ... More
Neurokinin B- and substance P-like immunoreactivity are co-localized in enteric nerves of rat ileum.
AuthorsYunker AM, Krause JE, Roth KA
JournalRegul Pept
PubMed ID10235636
The tachykinins (TKs) substance P (SP), neurokinin A (NKA), and neurokinin B (NKB) have conserved C-terminal sequences and mediate similar physiological responses by activating neurokinin receptors found on neural and smooth muscle cells. Many enteric nerves express preprotachykinin A (PPT A) mRNA and synthesize SP and NKA. However, it is ... More
An optimized method for in situ hybridization with signal amplification that allows the detection of rare mRNAs.
AuthorsYang H, Wanner IB, Roper SD, Chaudhari N
JournalJ Histochem Cytochem
PubMed ID10082745
In situ hybridization (ISH) using nonradioactive probes enables mRNAs to be detected with improved cell resolution but compromised sensitivity compared to ISH with radiolabeled probes. To detect rare mRNAs, we optimized several parameters for ISH using digoxygenin (DIG)-labeled probes, and adapted tyramide signal amplification (TSA) in combination with alkaline phosphatase ... More
Mice lacking link protein develop dwarfism and craniofacial abnormalities.
AuthorsWatanabe H, Yamada Y
JournalNat Genet
PubMed ID9988279
Link protein (LP), an extracellular matrix protein in cartilage, stabilizes aggregates of aggrecan and hyaluronan, giving cartilage its tensile strength and elasticity. Cartilage provides the template for endochondral ossification and is crucial for determining the length and width of the skeleton. During endochondral bone formation, hypertrophic chondrocytes die and the ... More
A highly sensitive immunofluorescence procedure for analyzing the subcellular distribution of GABAA receptor subunits in the human brain.
AuthorsLoup F, Weinmann O, Yonekawa Y, Aguzzi A, Wieser HG, Fritschy JM
JournalJ Histochem Cytochem
PubMed ID9742069
We designed a protocol to improve the immunohistochemical analysis of human brain structures, which overcomes the limited detection sensitivity, high background, and intense autofluorescence commonly associated with human tissue. This procedure was evaluated by using antibodies against major GABAA receptor subunits (alpha1, alpha2, alpha3, gamma2) in autopsy and surgical specimens. ... More
Biotin amplification of biotin and horseradish peroxidase signals in histochemical stains.
AuthorsAdams JC
JournalJ Histochem Cytochem
PubMed ID1527370
A procedure is described for intensifying histochemical reactions by amplification of biotinylated sites. This is achieved by deposition of biotinylated tyramine on the tissue through the enzymatic action of horseradish peroxidase (HRP). The amplified biotin sites are subsequently visualized by binding them to avidin, to which a marker is attached. ... More
Pre-embedding immunolabeling for electron microscopy: an evaluation of permeabilization methods and markers.
AuthorsHumbel BM, de Jong MD, Müller WH, Verkleij AJ
JournalMicrosc Res Tech
PubMed ID9712162
For scarce antigens or antigens which are embedded in a dense macromolecular structure, on-section labeling, the first method of choice, is not always successful. Often, the antigen can be localized by immunofluorescence microscopy, usually by a pre-embedding labeling method. Most of these methods lead to loss of ultrastructural details and, ... More
Detection of peroxisomal proteins and their mRNAs in serial sections of fetal and newborn mouse organs.
AuthorsGrabenbauer M, Fahimi HD, Baumgart E
JournalJ Histochem Cytochem
PubMed ID11156684
We present a protocol for detection of peroxisomal proteins and their corresponding mRNAs on consecutive serial sections of fetal and newborn mouse tissues by immunohistochemistry (IHC) and nonradioactive in situ hybridization (ISH). The use of perfusion-fixation with depolymerized paraformaldehyde combined with paraffin embedding and digoxigenin-labeled cRNA probes provided a highly ... More
Immunohistochemical signal amplification by catalyzed reporter deposition and its application in double immunostaining.
AuthorsHunyady B, Krempels K, Harta G, Mezey E
JournalJ Histochem Cytochem
PubMed ID8985127
The biotinyl-tyramide substrate of the horseradish peroxidase enzyme has been recently introduced to amplify immunohistochemical signals. We applied either fluorochromeor biotin-conjugated tyramine to improve the detection of different antigens in sections of rat stomach, pancreas, and hypothalamus. A ten- to 100-fold increase in staining efficiency was achieved, depending on the ... More
Human papillomavirus detection by non isotopic in situ hybridization, in situ hybridization with signal amplification and in situ polymerase chain reaction.
AuthorsBettinger D, Bernard B, Riethmuller D, Greslin I, Gay C, Lab M, Seillès E, Mougin C
JournalEur J Histochem
PubMed ID10563251
Classical in situ hybridization (ISH) with biotinylated probes makes it possible to detect and localize human papillomavirus (HPV) nucleic acid sequences in cytological and histological materials. This method is however of limited value in the detection of a few copies of the virus. Moreover the specificity of such a technique ... 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
Anterograde tracing and immunohistochemical characterization of potentially mechanosensitive vagal afferents in the esophagus.
AuthorsKressel M, Radespiel-Tröger M
JournalJ Comp Neurol
PubMed ID10440717
Vagal mechanosensitive afferents with an important functional role in esophageal peristalsis are well known from physiological studies. It is not known whether these fibers represent a separate subpopulation among all vagal afferents projecting to the esophageal wall. A morphological and immunohistochemical description of vagal afferents was undertaken to define their ... More
Application of catalyzed signal amplification in immunodetection of gonadotropin subunits in clinically nonfunctioning pituitary adenomas.
AuthorsSanno N, Teramoto A, Sugiyama M, Itoh Y, Osamura RY
JournalAm J Clin Pathol
PubMed ID8701926
Pituitary adenomas that are characterized by the absence of a particular clinical syndrome and the absence of excessive hormone secretion have been classified as nonfunctioning adenomas. Recent development of immunohistochemical analysis and hormonal assay have suggested that many of these tumors have function to secret the gonadotropin subunits. A novel ... More
Liver regeneration and alpha-fetoprotein messenger RNA expression in the retrorsine model for hepatocyte transplantation.
AuthorsDabeva MD, Laconi E, Oren R, Petkov PM, Hurston E, Shafritz DA
JournalCancer Res
PubMed ID9865742
Recently, we described a new model for hepatocyte transplantation with nearly total replacement of the liver by exogenous hepatocytes (E. Laconi et al., Am. J. Pathol., 153: 319-329, 1998). The model is based on the mitoinhibitory effect of the pyrrolizidine alkaloid retrorsine on hepatocytes in the resident liver while transplanted ... More
Localization of somatostatin receptors at the light and electron microscopial level by using antibodies raised against fusion proteins.
AuthorsHelboe L, Møller M
JournalProg Histochem Cytochem
PubMed ID10904940
Somatostatin mediates its multiple biological effects via specific plasma membrane receptors belonging to the family of G-protein coupled receptors with seven putative membrane-spanning domains. Five somatostatin receptor subtypes (sst1-sst5) have been cloned in human, mouse, and rat. We have raised specific antibodies against the five human somatostatin receptors by using ... More
Immunohistochemical distribution of the prohormone convertase PC5-A in rat brain.
AuthorsVilleneuve P, Seidah NG, Beaudet A
JournalNeuroscience
PubMed ID10408612
Prohormone convertase 5 is an endoprotease of the kexin/subtilisin-like family, which has been postulated to play a role in the proteolytic maturation of a variety of pro-peptides in the mammalian brain. In order to gain insight into the functional role of prohormone convertase 5 in the central nervous system, the ... More
ImmunoMax. A maximized immunohistochemical method for the retrieval and enhancement of hidden antigens.
BACKGROUND: Since the introduction of mAb, immunohistochemistry has become an important tool in research and in surgical pathology. The most widely used fixative in routine histopathology is formaldehyde, and it has become the gold standard for morphologic tissue preservation. Although the molecular mechanism underlying the tissue fixation is not well ... More
Sensitive mRNA detection by fluorescence in situ hybridization using horseradish peroxidase-labeled oligodeoxynucleotides and tyramide signal amplification.
Authorsvan de Corput MP, Dirks RW, van Gijlswijk RP, van Binnendijk E, Hattinger CM, de Paus RA, Landegent JE, Raap AK
JournalJ Histochem Cytochem
PubMed ID9774624
With the ongoing progress in human genome projects, many genes are discovered whose function and/or expression pattern are not known. Most of these genes are expressed in relatively low abundance compared to housekeeping genes such as elongation factor-1alpha and beta-actin. Gene expression is studied by Northern blot assays or by ... More
Primaquine interferes with membrane recycling from endosomes to the plasma membrane through a direct interaction with endosomes which does not involve neutralisation of endosomal pH nor osmotic swelling of endosomes.
Authorsvan Weert AW, Geuze HJ, Groothuis B, Stoorvogel W
JournalEur J Cell Biol
PubMed ID10928454
The anti-malaria drug primaquine is a weak base which accumulates in endosomes in a protonated form and consequently neutralises the endosomal pH. Bafilomycin A1 prevents endosome acidification by inhibiting the vacuolar proton pump. Although both agents neutralise the endosomal pH, only primaquine has a strong inhibitory effect on recycling of ... More
Extranuclear localization of endogenous 11beta-hydroxysteroid dehydrogenase-2 in aldosterone target cells.
AuthorsNáray-Fejes-Tóth A, Fejes-Tóth G
JournalEndocrinology
PubMed ID9607806
Type 2 11beta-hydroxysteroid dehydrogenase (11betaHSD2) plays a key role in conferring aldosterone selectivity on the mineralocorticoid receptor (MR) by inactivating intracellular glucocorticoids before they can occupy the MR. 11betaHSD2 is a microsomal enzyme expressed in aldosterone target cells, although its subcellular distribution is still unclear. The goal of this study ... 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
Op/op mice defective in production of functional colony-stimulating factor-1 lack macrophages in muscularis externa of the small intestine.
AuthorsMikkelsen HB, Thuneberg L
JournalCell Tissue Res
PubMed ID10022968
The osteopetrotic (op/op) mutant mouse possesses an inactivating mutation in the colony-stimulating factor-1 (CSF-1) gene, which results in the absence of certain macrophages and in osteopetrosis, following a lack of osteoclasts. Studies of the op/op mouse indicate that CSF-1-dependent tissue macrophages may belong to a trophic and/or scavenger subpopulation, which ... More
The use of MAB 1977 monoclonal antibody for the immunohistochemical localization of beta 1 integrins in paraffin-embedded human kidney.
AuthorsBernardini N, Bianchi F, Lupetti M, Dolfi A
JournalTumori
PubMed ID9267487
AIMS AND BACKGROUND: Integrins are widely known cell membrane receptors for extracellular matrix molecules. The beta 1 integrin subgroup is mainly expressed by kidney cells; immunolocalization of these molecules is usually carried out on cryostatic sections. A commercial monoclonal antibody directed against the human beta 1 integrin was tested in ... More