Fine Specificity Analysis with Monoclonal Antibodies of Antigens Controlled by the Major Histocompatibility Complex and by the Qa/TL Region in Mice

Lemke, Hilmar; Hämmerling, G. J.; Hämmerling, Ulrich

doi:10.1111/j.1600-065x.1979.tb00293.x

Cited by 421 publications

(123 citation statements)

References 30 publications

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“…The sections were pre-treated with peroxide (H 2 O 2 ) to quench endogenous peroxidase activity. MoAbs specifying the following cell surface markers were employed as primary reagents: CD11b [7] (Seralab, Crawley Down, UK), CD4 [8] and CD8 [9] (Becton Dickinson, Moutain View, CA), CD5 [10], CD25 [11] and CD45R (B220) (all from Pharmingen, San Diego, CA), and biotinylated mouse MoAbs to non-polymorphic murine MHC class II (clone 13/4) [12]. Rat MoAb to murine IL-2 (clone S4B6) [13] and interferon-gamma (IFN-) (clone XMG1.2) [14], gifts from Dr T. Mossman (Calgary, Canada), were used to detect the corresponding cytokine-containing cells.…”

Section: Immunohistochemical Techniquesmentioning

confidence: 99%

Experimental T cell-mediated inflammatory reactions in the murine oral mucosa. II. Immunohistochemical characterization of resident and infiltrating cells

Ahlfors

Jönsson²,

Czerkinsky

1996

Clinical and Experimental Immunology

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SUMMARYThe nature and phenotype of infiltrating cells in DTH-like reactions elicited in the murine oral mucosa have been examined by routine histological and immunohistochemical procedures. During the first few hours that followed buccal challenge with the contact sensitizer oxazolone, a discrete lymphocytic reaction was disclosed in the oral mucosa of animals previously sensitized at skin sites, but was absent in animals that had been sensitized at buccal sites. The early lymphocytic reaction in the oral mucosa of skin-sensitized animals preceded the emergence of CD11 + polymorphonuclear cells (PMN) which was most prominent 8 h after hapten challenge and invaded the whole thickness of the oral epithelium. The PMN rapidly disappeared by 24 h. In contrast, early PMN infiltration was virtually absent in specimens from animals similarly challenged but that had been sensitized at local buccal sites. Irrespective of site of initial sensitization, inflammatory reactions developed in the oral mucosa, being maximal by 24 h. At that stage, CD11 + macrophages were the predominant cell type. Both CD4 and CD8 T lymphocytes were scattered in both lamina propria and epithelium, and their numbers were raised throughout the time period studied (2-168 h). IL-2 receptor expression was maximal 16 h post-challenge, and was paralleled by increased DNA synthesis in CD4 + and CD8 + cells, as demonstrated by paired immunohistoautoradiography. Focal accumulations of mononuclear cells containing IL-2-producing cells were readily detected as early as 2-3 h following local challenge with hapten in animals primed at skin but not at buccal sites. Maximal IL-2 staining was detected at 24 h irrespective of initial sensitization site. Interferon-gamma-producing cells were detected at 8 h post-challenge and remained increased during the first 24 h. MHC class II expression was detected on few oral mucosa cells during the first 4 h following hapten challenge, being mainly confined to dendritic-like cells. Consistent with increased numbers of macrophages, MHC class II expression was most intense in specimens obtained 8-24 h after hapten challenge. Thereafter, MHC class II expression was still observed in specimens obtained as late as 72 h, but was essentially associated with patches of basal keratinocytes. Taken together, these observations support the notion that the murine oral mucosa can serve as the site of expression of locally or remotely induced DTH reactions, but also indicate that the site of initial sensitization can profoundly affect the cellular composition of inflammatory reactions subsequent to local buccal challenge.

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Section: Immunohistochemical Techniquesmentioning

confidence: 99%

Experimental T cell-mediated inflammatory reactions in the murine oral mucosa. II. Immunohistochemical characterization of resident and infiltrating cells

Ahlfors

Jönsson²,

Czerkinsky

1996

Clinical and Experimental Immunology

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“…This precipitation ofinsulin receptors by anti-H-2 antibodies could be due either to the presence of common epitopes on insulin receptors and H-2 antigens (cross-reactivity), or to the formation of a molecular complex between the two antigens (coprecipitation). The demonstration that different monoclonal antibodies (lanes B, D, and F), known to react with distinct antigenic determinants (18)(19)(20), are all able to precipitate the insulin receptor strongly supports the coprecipitation possibility.…”

mentioning

confidence: 84%

Molecular association between major histocompatibility complex class I antigens and insulin receptors in mouse liver membranes.

Fehlmann¹,

Peyron²,

Samson³

et al. 1985

Proc. Natl. Acad. Sci. U.S.A.

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RESULTS AND DISCUSSION Liver membranes were purified from congenic H-2k mice.Insulin receptors were specifically labeled with a 125I-labeled photoreactive insulin analogue, and membranes were solubilized in 1% Nonidet P-40. Immunoprecipitation of insulin receptors by anti-insulin receptor antibodies (17) (Fig. 1, lane A) revealed the presence of a major 130-kDa labeled polypeptide corresponding to the a subunit of the insulin receptor identified in a variety of tissues and cell types (21). Quantification of the radioactivity associated with the immunopreAbbreviations: MHC, major histocompatibility complex; EGF, epidermal growth factor. 8634The publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact.

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“…BALB/c and (C3H x DBA/2)F1 mice were obtained Monoclonal Antibodies and Staining Reagents We purified Kk-specific, monoclonal antibodies (mabs) from ascites fluid of BALB/c mice grafted with the respective hybridoma cell lines (13) by precipitation with 45% saturated ammonium sulfate and chromatography on DEAEcellulose (Whatman, Maidstone, England). The monoclonal antibodies are listed in Table 1.…”

Section: Cellsmentioning

confidence: 99%

Inexpensive upgrading of a FACS I and isolation of rare somatic variants by double‐fluorescence sorting

Weichel¹,

Liesegang²,

Gehrke³

et al. 1985

Cytometry

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We have modified a FACS I by addition of a tibodies labelled with FITC or Texas Red, we tunable dye laser and an optical system for have isolated from a murine T-cell lymphoma fluorescence detection that allows physically line variant subclones expressing structurally independent measurement of green and red altered histocompatibility class I (H-2Kk) immunofluorescence. These modifications are molecules. inexpensive and should be applicable to most single-laser systems. By using the modified machine and double-fluorescence activated Key words: Dual immunofluorescence, cell selection with H-2Kk-specific monoclonal an-sorting, Texas Red, H-2 variantsThe experiments reported in this paper were aimed at isolating structural variants from the mouse lymphoma line LDHB that express altered instead of wild type H2Kk molecules on the cell surface. These variants are interesting for the investigation of the structural basis of class I antigen function in immune recognition. Furthermore, they can be used to approach the question of which types of somatic mutation occur in the polymorphic family of class I genes.In our previous work, we had achieved this aim by inhibiting the binding of one fluorescent monoclonal anti-Kk antibody to the cells by an excess of a second, unlabelled one and selecting for rare fluorescent cells in the fluorescence activated cell sorter (FACS) (10). This approach was restricted to the selection of structural variants that had lost the expression of one of two Kk determinants, which had to be sufficiently close to each other (topographically) so that the binding of an antibody to one of them would inhibit the binding of a second antibody to the other. In the present work this limitation was overcome by staining wild type cells with two monoclonal anti-Kk antibodies that are labelled with green and red fluorescent tags, respectively, and enriching the cells that carry one label only. For this purpose, we have upgraded a single-laser FACS I instrument by addition of a dye laser that supplies the excitation light for red-fluorescent dyes, such as Texas Red or propidium iodide, that can be used for dual parameter sorting with fluorescein as a green fluorescent label. In contrast to other approaches to double (or triple) fluorescence (161, the dye laser is pumped by the same argon laser that provides the 488 nm light for fluorescein excitation. MATERIALS AND METHODS Cell SorterThe modifications described here were added to a FACS I instrument (Becton Dickinson, Mountain View, CA; serial no. 04). A 5-watt argon ion laser (SpectraPhysics Model 164-08, Darmstadt, FRG) operated at 4 Watts in an all-line mode to give approximately 800 milliwatts at 488 nm and 1.7 watt at 514 nm served as a source for 488 nm excitation of fluorescein and for pumping a Model 375 dye laser (Spectra-Physics, Darmstadt, FRG). The latter was operated with Rhodamin 6G Cambda Physik, Gottingen, FRG) in a setup as described by . Reflectors, dichroic mirrors, and interference and coloured-glass filters used in the system (Fig. 1)

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Fine Specificity Analysis with Monoclonal Antibodies of Antigens Controlled by the Major Histocompatibility Complex and by the Qa/TL Region in Mice

Cited by 421 publications

References 30 publications

Experimental T cell-mediated inflammatory reactions in the murine oral mucosa. II. Immunohistochemical characterization of resident and infiltrating cells

Experimental T cell-mediated inflammatory reactions in the murine oral mucosa. II. Immunohistochemical characterization of resident and infiltrating cells

Molecular association between major histocompatibility complex class I antigens and insulin receptors in mouse liver membranes.

Inexpensive upgrading of a FACS I and isolation of rare somatic variants by double‐fluorescence sorting

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