当前位置： SCI文献检索 > JOURNAL OF IMMUNOLOGICAL METHODS期刊下所有文献 > Reverse transcriptase in situ polymerase chain reaction for gene expression in rat mast cells and macrophages.

Reverse transcriptase in situ polymerase chain reaction for gene expression in rat mast cells and macrophages.

Abstract：

:Direct reverse transcriptase in situ polymerase chain reaction (RT-in situ PCR) of selected mRNA expression in rat mast cells (MC) and alveolar macrophages (AM) was optimized. Rat peritoneal mast cells (PMC), rat cultured mast cells (RCMC), rat bronchoalveolar lavage cells (BALC) or rat cultured alveolar macrophages (NR8383) were studied for the detection of mRNA for beta-actin, TNF-alpha and/or CD8alpha. Each type of cell has unique optimal conditions for RT-in situ PCR. The following parameters were carefully evaluated for optimization: protease digestion, DNAse digestion, heparinase digestion, RT, PCR cycle number and signal development with chromagen. Heparinase digestion was required for PMC mRNA detection because they contain large amounts of heparin proteoglycan, which is a potent inhibitor of RT and Taq polymerase enzymes. Only a few PCR cycles were needed to produce a cytoplasmic signal for mRNA transcripts in RCMC, whereas other types of cells (PMC, BALC and NR8383) needed at least 20 cycles for mRNA detection. The mRNA signal in PMC was localized to the perinuclear region, whereas mRNA in other cell types (RCMC, BALC and NR8383) were detected throughout the cytoplasm. Furthermore, modified Southern blot analysis for TNF-alpha in RCMC treated with RT-in situ PCR demonstrated the specificity of amplification product. The modified and optimized protocols for this procedure were successfully applied to detect and localize several mRNA transcripts in rat MC and AM. The approach is valuable and can be used to further study selected gene expression in these and other cell types.

journal_name

J Immunol Methods

journal_title

Journal of immunological methods

authors

Nohara O,Gilchrist M,Déry RE,Stenton GR,Hirji NS,Befus AD

doi

10.1016/s0022-1759(99)00061-7

keywords:

subject

Has Abstract

pub_date

1999-06-24 00:00:00

pages

147-58

issue

1-2

eissn

0022-1759

issn

1872-7905

pii

S0022-1759(99)00061-7

journal_volume

226

pub_type

杂志文章

相关文献

JOURNAL OF IMMUNOLOGICAL METHODS文献大全
  • Monopalmitic acid-peptide conjugates induce cytotoxic T cell responses against malarial epitopes: importance of spacer amino acids.

    abstract：:Cytolytic T cells (CTL) play a critical role in providing protection against the liver stage of malaria infection. Previous investigations have shown that induction of CTL against peptide or proteins can be achieved by attachment of lipids. In the present study, we used the Plasmodium berghei circumsporozoite protein ...

    journal_title：Journal of immunological methods

    pub_type： 杂志文章

    doi：10.1016/0022-1759(95)00052-c

    authors： Verheul AF,Udhayakumar V,Jue DL,Wohlhueter RM,Lal AA

    更新日期：1995-06-09 00:00:00

  • Rapid single-step FCGR3A genotyping based on SYBR Green I fluorescence in real-time multiplex allele-specific PCR.

    abstract：:A polymorphism at position 559 in the cDNA of the FCGR3A gene encoding the FcgammaRIIIa generates two allotypes with either a valine (Val) or a phenylalanine (Phe) at amino acid position 158. This polymorphism is of major importance in immunopathology and in pharmacogenetics, especially monoclonal antibody treatments....

    journal_title：Journal of immunological methods

    pub_type： 杂志文章

    doi：10.1016/s0022-1759(03)00123-6

    authors： Dall'Ozzo S,Andres C,Bardos P,Watier H,Thibault G

    更新日期：2003-06-01 00:00:00

  • Fluorometric enzyme immunoassay for measurement of infectious feline leukemia virus and its neutralization.

    abstract：:A fluorometric enzyme immunoassay using an alkaline phosphatase-conjugated monoclonal antibody was developed to quantitate feline leukemia virus (FeLV) infection. Monoclonal antibodies, directed against the FeLV structural protein p27, were conjugated with alkaline phosphatase using a modified maleimide method. The en...

    journal_title：Journal of immunological methods

    pub_type： 杂志文章

    doi：10.1016/0022-1759(88)90181-0

    authors： Kushner NN,Riggin CH,Annunziato ME,Marciani DJ

    更新日期：1988-11-10 00:00:00

  • Characterization of antibodies to advanced glycosylation end products on protein.

    abstract：:Antibodies directed against advanced glycosylation end products (AGEs) formed during a Maillard reaction have been generated and characterized. Since protein-bound AGEs recognized by the antibodies were labile to acid hydrolysis, the antibodies were further characterized by using the AGE-alpha-acetyl-L-lysine methyl e...

    journal_title：Journal of immunological methods

    pub_type： 杂志文章

    doi：10.1016/0022-1759(91)90133-z

    authors： Nakayama H,Taneda S,Mitsuhashi T,Kuwajima S,Aoki S,Kuroda Y,Misawa K,Yanagisawa K,Nakagawa S

    更新日期：1991-06-24 00:00:00

  • In situ hybridization for cytokine mRNA with digoxigenin-labeled riboprobes. Sensitivity of detection and double label applications.

    abstract：:A sensitive in situ hybridization procedure using both digoxigenin and 35S-labeled riboprobes is described that allows detection of single T cells expressing cytokine mRNA species in both single and double label formats. Modifications to existing procedures have been developed that allow in situ hybridization to be pe...

    journal_title：Journal of immunological methods

    pub_type： 杂志文章

    doi：10.1016/0022-1759(95)00027-8

    authors： Karr LJ,Panoskaltsis-Mortari A,Li J,Devore-Carter D,Weaver CT,Bucy RP

    更新日期：1995-05-11 00:00:00

  • Rapid dissociation of adherent human tumor cells by ultrasound.

    abstract：:Cultured human melanoma and gastrointestinal carcinoma cells were detached from substrate and further dissociated by placing the culture vessel into a water-filled ultrasonic cleaner (43 kHz) and sonicating it for 10-50 s. Plating efficiency and long-term growth of three melanoma cell lines were similar after ultrasou...

    journal_title：Journal of immunological methods

    pub_type： 杂志文章

    doi：10.1016/0022-1759(87)90480-7

    authors： Menssen HD,Herlyn M,Rodeck U,Koprowski H

    更新日期：1987-11-23 00:00:00

  • Characterisation of monoclonal antibodies against human interleukin-10 and their use in an ELISA for the measurement of this cytokine.

    abstract：:Biological and biochemical characteristics of monoclonal antibodies (MABs) raised against human interleukin-10 (IL-10) are described as well as their use in the design of a specific ELISA for the measurement of the cytokine. 21 murine anti-human interleukin-10 (IL-10) MABs were obtained by fusion of splenocytes from m...

    journal_title：Journal of immunological methods

    pub_type： 杂志文章

    doi：10.1016/0022-1759(94)90160-0

    authors： De Groote D,Marchant A,Fauchet F,Jadoul M,Dehart I,Gérard C,Gevaert Y,Lopez M,Gathy R,Franssen JD

    更新日期：1994-12-28 00:00:00

  • From big flow cytometry datasets to smart diagnostic strategies: The EuroFlow approach.

    abstract：:The rise in the analytical speed of mutiparameter flow cytometers made possible by the introduction of digital instruments, has brought up the possibility to manage progressively higher number of parameters simultaneously on significantly greater numbers of individual cells. This has led to an exponential increase in ...

    journal_title：Journal of immunological methods

    pub_type： 杂志文章,评审

    doi：10.1016/j.jim.2019.07.003

    authors： Pedreira CE,Costa ESD,Lecrevise Q,Grigore G,Fluxa R,Verde J,Hernandez J,van Dongen JJM,Orfao A,EuroFlow.

    更新日期：2019-12-01 00:00:00

  • An antigen-specific, four-color, B-cell FluoroSpot assay utilizing tagged antigens for detection.

    abstract：:The FluoroSpot assay, a variant of ELISpot utilizing fluorescent detection, has so far been used primarily for assessment of T cells, where simultaneous detection of several cytokines has allowed a more qualitative analysis of functionally distinct T cells. The potential to measure multiple analytes also presents seve...

    journal_title：Journal of immunological methods

    pub_type： 杂志文章

    doi：10.1016/j.jim.2016.02.020

    authors： Jahnmatz P,Bengtsson T,Zuber B,Färnert A,Ahlborg N

    更新日期：2016-06-01 00:00:00

  • Dilatometry, a neglected method in immunological studies.

    abstract：:Dilatometry has become a useful method for the study of proteins owing to its simplicity and accuracy. However, it has seldom been used in the study of immunoglobulins. Therefore possible applications of the method for that study are being discussed. A description is also given of the most common experimental set-up a...

    journal_title：Journal of immunological methods

    pub_type： 杂志文章,评审

    doi：10.1016/0022-1759(75)90110-6

    authors： Lapanje S,Skerjanc J

    更新日期：1975-12-01 00:00:00

  • CD19+ B lymphocytes are the major source of human antibody-secreting hybridomas generated by electrofusion.

    abstract：:Human monoclonal antibodies may be generated by electrofusion of human B lymphocytes with a human/mouse heteromyeloma line. In addition to a fusion protocol optimised for the fusion partners, the activation of B lymphocytes is crucial for fusion and hybrid efficiency. In this study, we initially treated peripheral blo...

    journal_title：Journal of immunological methods

    pub_type： 杂志文章

    doi：10.1016/s0022-1759(01)00431-8

    authors： Schmidt E,Leinfelder U,Gessner P,Zillikens D,Bröcker EB,Zimmermann U

    更新日期：2001-09-01 00:00:00

  • Immunocytochemical detection of 5-bromo-2-deoxyuridine incorporation in individual cells.

    abstract：:5-Bromo-2-deoxyuridine (BrdUrd) is a thymidine analogue which can be used to detect DNA synthesis in cells. The present paper describes a method for the fixation and DNA denaturation of cells that affects neither the morphology of the cells not their adhesion to a glass surface. The cells were fixed in absolute ethano...

    journal_title：Journal of immunological methods

    pub_type： 杂志文章

    doi：10.1016/0022-1759(88)90401-2

    authors： Van Furth R,Van Zwet TL

    更新日期：1988-04-06 00:00:00

  • The use of targetted liposomes to isolate cells bearing immunoglobulin receptors.

    abstract：:A new method of mouse B cell isolation has been developed. It is based on the differences in buoyant density of free lymphocytes and cells bound within liposome-cellular complexes (LCC). LCC arise during the interaction of B cells bearing surface immunoglobulins (Ig) with liposomes loaded with affinity purified rabbit...

    journal_title：Journal of immunological methods

    pub_type： 杂志文章

    doi：10.1016/0022-1759(89)90074-4

    authors： Golubev A,Sidorova E

    更新日期：1989-12-20 00:00:00

  • The identification of the subclasses of human IgG by analytical isotachophoresis.

    abstract：:A new method is described for demonstrating the individual subclasses of human IgG immunoglobulins by means of analytical isotachophoresis. Individual subclasses of IgG purified from sera of multiple myeloma patients have distinct mobility patterns. When a mixture with purified IgG for each of the 4 subclasses is anal...

    journal_title：Journal of immunological methods

    pub_type： 杂志文章

    doi：10.1016/0022-1759(79)90163-7

    authors： Hedlund KW,Wistar R Jr,Nichelson D

    更新日期：1979-01-01 00:00:00

  • Inhibition of clone formation as an assay for T cell-mediated cytotoxicity: short-term kinetics and comparison with 51Cr release.

    abstract：:The short-term kinetics of T cell-mediated cytotoxicity was investigated using a cloning inhibition assay. Murine cytotoxic thymus-derived lymphocytes (CTL) generated in vitro in mixed leukocyte cultures (MLC), were incubated for various periods of time at 37 degrees C with allogeneic mastocytoma target cells. The mix...

    journal_title：Journal of immunological methods

    pub_type： 杂志文章

    doi：10.1016/0022-1759(77)90201-0

    authors： Lees RK,MacDonald HR,Sinclair NR

    更新日期：1977-01-01 00:00:00

  • A software package for the calculation and statistical analysis of 51Cr-release assays.

    abstract：:A software package has been developed to calculate, store, and statistically analyze the results of 51Cr-release assays. It is written in Applesoft BASIC, is 'user friendly', and contains error trapping routines to catch most common mistakes. The greeting program contains a master menu from which one of the following ...

    journal_title：Journal of immunological methods

    pub_type： 杂志文章

    doi：10.1016/0022-1759(84)90004-8

    authors： Hogan KT,Evans DL

    更新日期：1984-09-04 00:00:00

  • Characterization of the diversity of T cell receptor γδ complementary determinant region 3 in human peripheral blood by Immune Repertoire Sequencing.

    abstract：:γδ T cells function as sentinels in early host response to infections and malignancies. Although γδ T cells are regarded as innate immune cells and recognize antigens in a non-MHC restricted manner, they possess a huge diversity of complementary determinant region 3 (CDR3) of T cell receptor (TCR) generated by the rea...

    journal_title：Journal of immunological methods

    pub_type： 杂志文章

    doi：10.1016/j.jim.2017.01.009

    authors： Chen H,Zou M,Teng D,Zhang J,He W

    更新日期：2017-04-01 00:00:00

  • A hemolytic plaque assay for the detection of red blood cells carrying abnormal or mutant hemoglobin.

    abstract：:A simple hemolytic plaque assay for the detection of red blood cells carrying abnormal or mutant hemoglobin is described. The assay is based on the use, as indicator cells, of sheep red blood cells coated with monospecific antibody against the hemoglobin variant, another complement fixing multispecific anti-hemoglobin...

    journal_title：Journal of immunological methods

    pub_type： 杂志文章

    doi：10.1016/0022-1759(82)90075-8

    authors： Baig MA,Ansari AA,Malling HV

    更新日期：1982-11-26 00:00:00

  • Analysis of the effector activity of Ara h 2 and Ara h 6 by selective depletion from a crude peanut extract.

    abstract：:It is important to know the contribution of specific allergens to a complex allergenic extract and to have a dependable method to assess the effector activity of an extract specifically depleted of that allergen. We have previously shown that removal of the major peanut allergen, Ara h 2, from a crude peanut extract (...

    journal_title：Journal of immunological methods

    pub_type： 杂志文章

    doi：10.1016/j.jim.2011.06.031

    authors： Chen X,Zhuang Y,Wang Q,Moutsoglou D,Ruiz G,Yen SE,Dreskin SC

    更新日期：2011-09-30 00:00:00

  • Protein transduction as a means of effective manipulation of Cdc42 activity in primary T cells.

    abstract：:The Rho family GTPase Cdc42 is a critical regulator of cellular polarization from yeast to man. An analysis of its function in T cell activation is therefore of interest. This analysis poses two substantial challenges, similar to the analysis of many other critical T cell signaling intermediates. First, Cdc42 is requi...

    journal_title：Journal of immunological methods

    pub_type： 杂志文章

    doi：10.1016/j.jim.2006.10.017

    authors： Tskvitaria-Fuller I,Mistry N,Sun S,Wülfing C

    更新日期：2007-01-30 00:00:00

  • An efficient protocol for in vivo labeling of proliferating epithelial cells.

    abstract：:The study of organogenesis, tissue-homeostasis and regeneration requires the precise assessment of in vivo cell proliferation. To this end a host of methods have been developed to detect and quantify DNA synthesis in proliferating cells. These include cell labeling with various nucleotide analogues and fluorescence re...

    journal_title：Journal of immunological methods

    pub_type： 杂志文章

    doi：10.1016/j.jim.2018.03.015

    authors： Michel C,Küchler R,Martin I,Kyewski B,Pinto S

    更新日期：2018-06-01 00:00:00

  • The demonstration of cell surface antigens on T cells, B cells and accessory cells in paraffin-embedded human tissues.

    abstract：:This paper describes a method for processing fresh tissue that allows immunohistological analysis on paraffin sections. The method is based on the use of periodate-lysine-paraformaldehyde fixation. The effects of variation in fixation time, concentration of paraformaldehyde, dehydration, clearing, wax embedding and en...

    journal_title：Journal of immunological methods

    pub_type： 杂志文章

    doi：10.1016/0022-1759(84)90106-6

    authors： Collings LA,Poulter LW,Janossy G

    更新日期：1984-12-31 00:00:00

  • A sensitive proliferation assay to determine the specific T cell response against HLA-A2.1-binding peptides.

    abstract：:A proliferation assay to detect a specific T cell response against HLA class I-binding peptides is reported. To establish the specificity and sensitivity of the assay we used a synthetic peptide derived from the CMV glycoprotein B containing the HLA-A2.1 ligand motif. Lymphocyte proliferation was measured following cu...

    journal_title：Journal of immunological methods

    pub_type： 杂志文章

    doi：10.1016/0022-1759(96)00124-x

    authors： Mayer S,Scheibenbogen C,Lee KH,Keilholz W,Stevanovic S,Rammensee HG,Keilholz U

    更新日期：1996-10-16 00:00:00

  • In vivo 6-thioguanine-resistant T cells from melanoma patients have public TCR and share TCR beta amino acid sequences with melanoma-reactive T cells.

    abstract：:In vivo hypoxanthine-guanine phosphoribosyltransferase (HPRT)-deficient T cells (MT) from melanoma patients are enriched for T cells with in vivo clonal amplifications that traffic between blood and tumor tissues. Melanoma is thus a model cancer to test the hypothesis that in vivo MT from cancer patients can be used a...

    journal_title：Journal of immunological methods

    pub_type： 杂志文章

    doi：10.1016/j.jim.2010.12.007

    authors： Zuleger CL,Macklin MD,Bostwick BL,Pei Q,Newton MA,Albertini MR

    更新日期：2011-02-28 00:00:00

  • Dinitrophenyl (DNP) hapten sandwich staining (DHSS) procedure. A 10 year review of its principle reagents and applications.

    abstract：:Over the past 10 years an immunoperoxidase method using dinitrophenyl (DNP) hapten-labelled primary or secondary probes has been devised. Its widely successful application in research and diagnostic work has depended upon the development of certain key reagents. These include a novel non-deleterious DNP labelling comp...

    journal_title：Journal of immunological methods

    pub_type： 杂志文章,评审

    doi：10.1016/0022-1759(92)90078-8

    authors： Jasani B,Thomas ND,Navabi H,Millar DM,Newman GR,Gee J,Williams ED

    更新日期：1992-06-24 00:00:00

  • Optimization of the sulforhodamine B colorimetric assay.

    abstract：:Sulforhodamine B (SRB) protein staining has been widely used for cell proliferation and chemosensitivity testing, substituting for tetrazolium-based assays. However, the cell fixation step in the original assay is subject to error. We tested whether aspiration of medium with an automatic microplate multiwash device pr...

    journal_title：Journal of immunological methods

    pub_type： 杂志文章

    doi：10.1016/s0022-1759(97)00137-3

    authors： Papazisis KT,Geromichalos GD,Dimitriadis KA,Kortsaris AH

    更新日期：1997-10-27 00:00:00

  • The use of T bag synthesis with paper discs as the solid phase in epitope mapping studies.

    abstract：:Epitope mapping with synthetic peptides bound to derivatized paper discs has been investigated using the discs both as a solid-phase matrix for peptide synthesis as well as the solid phase in immunologic testing procedures without detachment of the peptides from the paper discs. Using the T bag (tea bag) method the si...

    journal_title：Journal of immunological methods

    pub_type： 杂志文章

    doi：10.1016/0022-1759(93)90293-g

    authors： van 't Hof W,van den Berg M,Aalberse RC

    更新日期：1993-05-26 00:00:00

  • An enzyme-linked immunosorbent assay (ELISA) for the detection of IgG and IgM anti-idiotypes directed against anti-HBs molecules.

    abstract：:A simple and specific enzyme-linked immunosorbent assay (ELISA) has been developed to detect circulating IgG and IgM anti-idiotypic antibodies directed against anti-HBs molecules using 96-well polyvinyl microtitre plates as the solid phase and HRPO-labelled goat anti-HBs as conjugate. Anti-idiotype reactions were obse...

    journal_title：Journal of immunological methods

    pub_type： 杂志文章

    doi：10.1016/0022-1759(87)90316-4

    authors： Irshad M,Gandhi BM,Acharya SK,Joshi YK,Tandon BN

    更新日期：1987-02-11 00:00:00

  • A europium fluoroimmunoassay for measuring binding of antigen to class II MHC glycoproteins.

    abstract：:A dissociation-enhanced lanthanide fluoroimmunoassay employing europium-streptavidin and time-resolved fluorimetry was developed to measure binding of biotin-labeled peptides to class II MHC proteins. Binding of biotin-peptides as measured by this assay was saturable and inhibited in the presence of unlabeled peptide....

    journal_title：Journal of immunological methods

    pub_type： 杂志文章

    doi：10.1016/0022-1759(93)90124-p

    authors： Tompkins SM,Rota PA,Moore JC,Jensen PE

    更新日期：1993-08-09 00:00:00

  • A method for the isolation and purification of mouse peripheral blood monocytes.

    abstract：:Studies in mice have advanced our understanding of the contribution of monocytes to inflammation and immunity. However, the lack of a straightforward and reliable method for the isolation of mouse blood monocytes remains a hurdle. Here we describe a fast and easy method for isolating monocytes from mouse blood based o...

    journal_title：Journal of immunological methods

    pub_type： 杂志文章

    doi：10.1016/j.jim.2010.04.004

    authors： Houthuys E,Movahedi K,De Baetselier P,Van Ginderachter JA,Brouckaert P

    更新日期：2010-07-31 00:00:00