scholarly journals Characterization of MAX.3 antigen, a glycoprotein expressed on mature macrophages, dendritic cells and blood platelets: identity with CD84

2000 ◽  
Vol 346 (3) ◽  
pp. 729-736 ◽  
Author(s):  
Stefan W. KRAUSE ◽  
Michael REHLI ◽  
Sven HEINZ ◽  
Reinhard EBNER ◽  
Reinhard ANDREESEN
Keyword(s):  
Dendritic Cells ◽  
Blood Platelets ◽  
Cell Types ◽  
Variable Expression ◽  
Surface Molecules ◽  
Sds Page ◽  
Molecular Masses ◽  
Antigen A

MAX.3 is a monoclonal antibody that preferentially reacts with mature macrophages (MAC), monocyte-derived dendritic cells, megakaryocytes and platelets. In this study, we describe the characterization, purification and identification of the MAX.3 antigen. Immunoprecipitation and SDS/PAGE revealed different molecular masses of MAX.3 antigen in MAC (60-90 kDa) and platelets (58-64 kDa), whereas a similar size (45 kDa) was observed in both cell types after digestion with N-glycosidase F. Lectin affinity and sequential treatment with different glycosidases suggests complex type glycosylation of MAX.3 antigen in MAC and hybrid type glycosylation in platelets. Amino acid sequencing led to the identification of a corresponding cDNA clone and showed its identity to the sequence of the CD84 antigen, a member of the CD2 family of cell surface molecules. MAX.3/CD84 was further studied by immunohistochemistry and a variable expression was found on tissue MAC, confirming this antigen to be mainly a marker for MAC in situ.

scholarly journals Characterization of MAX.3 antigen, a glycoprotein expressed on mature macrophages, dendritic cells and blood platelets: identity with CD84

2000 ◽  
Vol 346 (3) ◽  
pp. 729 ◽  
Author(s):  
Stefan W. KRAUSE ◽  
Michael REHLI ◽  
Sven HEINZ ◽  
Reinhard EBNER ◽  
Reinhard ANDREESEN
Keyword(s):  
Dendritic Cells ◽  
Blood Platelets ◽  
Antigen A

scholarly journals Exo-enzymatic addition of diazirine-modified sialic acid to cell surfaces enables photocrosslinking of glycoproteins

2021 ◽  
Author(s):  
Nageswari Yarravarapu ◽  
Rohit Sai Reddy Konada ◽  
Narek Darabedian ◽  
Nichole J. Pedowtiz ◽  
Soumya N. Krishnamurthy ◽  
...  
Keyword(s):  
Sialic Acid ◽  
Cell Surface ◽  
Cell Types ◽  
Cell Surfaces ◽  
Binding Interactions ◽  
Multiple Cell ◽  
Labeling Method ◽  
Cell Surface Glycoconjugates

Glycan binding often mediates extracellular macromolecular recognition events. Accurate characterization of these binding interactions can be difficult because of dissociation and scrambling that occur during purification and analysis steps. Use of photocrosslinking methods has been pursued to covalently capture glycan-dependent interactions in situ however use of metabolic glycan engineering methods to incorporate photocrosslinking sugar analogs is limited to certain cell types. Here we report an exo-enzymatic labeling method to add a diazirine-modified sialic acid (SiaDAz) to cell surface glycoconjugates. The method involves chemoenzymatic synthesis of diazirine-modified CMP-sialic acid (CMP-SiaDAz), followed by sialyltransferase-catalyzed addition of SiaDAz to desialylated cell surfaces. Cell surface SiaDAz-ylation is compatible with multiple cell types and is facilitated by endogenous extracellular sialyltransferase activity present in Daudi B cells. This method for extracellular addition of α2-6-linked SiaDAz enables UV-induced crosslinking of CD22, demonstrating the utility for covalent capture of glycan-mediated binding interactions.

scholarly journals Distribution of horseradish peroxidase (HRP)-anti-HRP immune complexes in mouse spleen with special reference to follicular dendritic cells.

1978 ◽  
Vol 79 (1) ◽  
pp. 184-199 ◽  
Author(s):  
L L Chen ◽  
A M Frank ◽  
J C Adams ◽  
R M Steinman
Keyword(s):  
Dendritic Cells ◽  
Horseradish Peroxidase ◽  
Immune Complexes ◽  
Cell Types ◽  
Mouse Spleen ◽  
Follicular Dendritic Cells ◽  
Quantitative Studies ◽  
Wide Range ◽  
Follicular Dendritic

The distribution of immune complexes has been studied in mouse spleen stimulated to contain many germinal centers (GC's). Horseradish peroxidase (HRP)-anti-HRP complexes were used as an appropriately precise and sensitive model. We were primarily interested in the relative abilities of three cell types to interact with complexes: lymphocytes, macrophages, and follicular dendritic cells (FDC's). The latter are distinctive, nonendocytic, stellate cells located primarily at the transition of mantle and GC zones of 2 degrees lymphoid follicles (Chen, L. L., J. C. Adams, and R. M. Steinman, 1978, J. Cell Biol. 77:148). Binding of immune complexes to lymphocytes could not be visualized in situ. Macrophages avidly interiorized complexes into lysosomes, but did not retain them extracellularly. In contrast, FDC's could retain HRP-anti-HRP extracellularly under appropriate conditions, but did not endocytose them. Cytochemical reactivity accumulated progressively on FDC's 1--6 h after administration of complexes i.v., remained stable in amount and location for 1 day, and then was progressively lost over a 1- to 5-day period. Several variables in the association of complexes with macrophages and FDC's were pursued. Only 1 microgram of complexed HRP had to be administered to visualize binding to both cell types. Macrophages interiorized complexes formed in a wide range of HRP/anti-HRP ratios, while FDC's associated with complexes formed in HRP excess only. Quantitative studies with [125I]HRP-anti-HRP demonstrated that 20% of the splenic load of HRP associated with FDC's. Complexes formed with an F(ab')2 anti-HRP were distributed primarily in macrophages. When the levels of the third component of serum complement were depleted by prior treatment with cobra venom factor, uptake of complexes by macrophages was reduced some 50% whereas association with FDC's was abolished. The fact that antigen excess complexes are retained extracellularly strengthens the idea that they are immunogenic. Finally, the association of complexes with FDC's seems to retard the entry of antigen into the GC proper.

Electrophoretic characterization of endo-(1,4)-β-glucanases secreted during assimilative growth and antheridiol-induced branching inAchlya ambisexualis

1996 ◽  
Vol 42 (6) ◽  
pp. 557-561 ◽  
Author(s):  
Terry W. Hill
Keyword(s):  
Enzyme Activity ◽  
Sodium Dodecyl Sulfate ◽  
Cell Walls ◽  
Sodium Dodecyl ◽  
Achlya Ambisexualis ◽  
Dodecyl Sulfate ◽  
Exclusion Chromatography ◽  
Molecular Masses

Secreted endo-(1,4)-β-glucanases ("cellulases") of Achlya ambisexualis were analyzed by a technique that permits visualization of enzyme activity in situ after electrophoresis in gels containing sodium dodecyl sulfate. Catalytic polypeptides with molecular masses of about 97, 74, 36, 29, and 25 kDa were observed in media from young cultures, though progressively fewer bands were observed as cultures aged. Based on size estimations of native enzymes with gel exclusion chromatography, the 97- and 36-kDa polypeptides were concluded to be subunits of a 245-kDa holoenzyme and the 25-kDa polypeptides were concluded to be subunits of a second holoenzyme of about 92 kDa. The data were insufficient to allow similar assignments for the more ephemeral 74- and 29-kDa polypeptides. The endoglucanases secreted during branch induction by antheridiol or 0.2% peptone comigrated in electrophoretic gels with enzymes secreted during normal assimilative growth. No endoglucanases specific to induced branching were observed.Key words: oomycetes, cell walls, endoglucanases, cellulases, antheridiol.

scholarly journals The mt1 Melatonin Receptor and RORβ Receptor Are Co-localized in Specific TSH-immunoreactive Cells in the Pars Tuberalis of the Rat Pituitary

2002 ◽  
Vol 50 (12) ◽  
pp. 1647-1657 ◽  
Author(s):  
Paul Klosen ◽  
Christele Bienvenu ◽  
Olivier Demarteau ◽  
Hugues Dardente ◽  
Hilda Guerrero ◽  
...  
Keyword(s):  
Cell Types ◽  
Orphan Receptor ◽  
Seasonal Effects ◽  
Pars Tuberalis ◽  
Melatonin Receptor ◽  
Functional Studies ◽  
Rat Pituitary ◽  
Mt1 Melatonin Receptor

The pars tuberalis (PT) of the pituitary represents an important target site for the time-pacing pineal hormone melatonin because it expresses a large number of mt1 receptors. Functional studies suggest that the PT mediates the seasonal effects of melatonin on prolactin (PRL) secretion. The aim of this study was the characterization of the pheno-type of melatonin-responsive cells. Furthermore, we determined whether RORβ, a retinoid orphan receptor present in the PT, was co-expressed in the same cells. We combined nonradioactive in situ hybridization (ISH) with hapten-labeled riboprobes for detection of the receptors and immunocytochemistry (ICC) for detection of αGSU (α-glycoprotein subunit), βTSH, βFSH, βLH, GH, PRL, and ACTH. Expression of mt1 mRNA was found in small round cells, co-localized with αGSU and βTSH. However, not all βTSH-containing cells expressed mt1 mRNA. The distribution of mt1- and RORβ-positive cells appeared to overlap, although more cells were labeled for RORβ than for mt1. Gonadotrophs, as well as other pars distalis cell types, were never labeled for mt1 melatonin receptor. Therefore, this study identifies the “specific” cells of the PT as the mt1 melatonin receptor-expressing cells.

scholarly journals Human Tumor-Infiltrating Dendritic Cells: From in Situ Visualization to High-Dimensional Analyses

Cancers ◽  
2019 ◽  
Vol 11 (8) ◽  
pp. 1082 ◽  
Author(s):  
Hubert ◽  
Gobbini ◽  
Bendriss-Vermare ◽  
Caux ◽  
Valladeau-Guilemond
Keyword(s):  
Dendritic Cells ◽  
T Cell Activation ◽  
Cell Activation ◽  
Human Tumor ◽  
High Dimensional ◽  
Prognostic Impact ◽  
In Situ Visualization ◽  
A Cell

The interaction between tumor cells and the immune system is considered to be a dynamic process. Dendritic cells (DCs) play a pivotal role in anti-tumor immunity owing to their outstanding T cell activation ability. Their functions and activities are broad ranged, triggering different mechanisms and responses to the DC subset. Several studies identified in situ human tumor-infiltrating DCs by immunostaining using a limited number of markers. However, considering the heterogeneity of DC subsets, the identification of each subtype present in the immune infiltrate is essential. To achieve this, studies initially relied on flow cytometry analyses to provide a precise characterization of tumor-associated DC subsets based on a combination of multiple markers. The concomitant development of advanced technologies, such as mass cytometry or complete transcriptome sequencing of a cell population or at a single cell level, has provided further details on previously identified populations, has unveiled previously unknown populations, and has finally led to the standardization of the DCs classification across tissues and species. Here, we review the evolution of tumor-associated DC description, from in situ visualization to their characterization with high-dimensional technologies, and the clinical use of these findings specifically focusing on the prognostic impact of DCs in cancers.

scholarly journals Partial Characterization of a Vicilin-Like Glycoprotein from Seeds of Flowering Tobacco (Nicotiana sylvestris)

2009 ◽  
Vol 2009 ◽  
pp. 1-12 ◽  
Author(s):  
Jared Q. Gerlach ◽  
Veer P. Bhavanandan ◽  
Paul A. Haynes ◽  
Lokesh Joshi
Keyword(s):  
Compositional Analysis ◽  
Nicotiana Sylvestris ◽  
Proteolytic Processing ◽  
Protein Subunits ◽  
Lectin Staining ◽  
Disulphide Bonds ◽  
Sds Page ◽  
Solanaceae Family

A vicilin-like glycoprotein from the seeds of Nicotiana sylvestris, flowering tobacco, has been identified using nanoLC/ESI-MS/MS. Sequences from a fragment of protein demonstrated homology with vicilins from other members of the Solanaceae family, notably potato (Solanum demissum). Reducing and nonreducing SDS-PAGE analyses of the identified protein indicated that fragments resulting from in situ proteolytic processing are joined by intrachain disulphide bonds. Staining with Con A lectin was specifically inhibited by mannose suggested the presence of -linked glycosylation which was confirmed by carbohydrate compositional analysis of PVDF-bound protein subunits. HPAEC-PAD analysis of the monosaccharides released from the glycoprotein by acid hydrolysis revealed glucosamine and mannose. -acetylglucosamine termination of attached oligosaccharides was further verified by inhibitable WGA lectin staining. Immunostaining of PVDF-bound N. sylvestris proteins with antibodies against G. max total protein demonstrated cross-staining at masses corresponding to fragments from the proteolytically processed protein subunits.

scholarly journals Purification and characterization of three extracellular (1→3)-β-d-glucan glucohydrolases from the filamentous fungus Acremonium persicinum

1995 ◽  
Vol 308 (3) ◽  
pp. 733-741 ◽  
Author(s):  
S M Pitson ◽  
R J Seviour ◽  
B M McDougall ◽  
J R Woodward ◽  
B A Stone
Keyword(s):  
Filamentous Fungus ◽  
Gel Filtration ◽  
High Specificity ◽  
Major Product ◽  
Ph Optimum ◽  
Sds Page ◽  
Molecular Masses ◽  
Monomeric Proteins ◽  
Ph Range

Three (1-->3)-beta-D-glucanases (GNs) were isolated from the culture filtrates of the filamentous fungus Acremonium persicinum and purified by (NH4)2SO4 precipitation followed by anion-exchange and gel-filtration chromatography. Homogeneity of the purified proteins was confirmed by SDS/PAGE, isoelectric focusing and N-terminal amino acid sequencing. All three GNs (GN I, II and III) are non-glycosylated, monomeric proteins with apparent molecular masses, estimated by SDS/PAGE, of 81, 85 and 89 kDa respectively. pI values for the three enzymes are 5.3, 5.1, and 4.4 respectively. The pH optimum for GN I is 6.5, and 5.0 for GN II and III. All three purified enzymes displayed stability over the pH range 4.5-10.0. Optimum activities for GN I, II and III were recorded at 65, 55 and 60 degrees C respectively, with both GN II and III having short-term stability up to 50 degrees C and GN I up to 55 degrees C. The purified GNs have high specificity for (1-->3)-beta-linkages and hydrolysed a range of (1-->3)-beta- and (1-->3)(1-->6)-beta-D-glucans, with laminarin from Laminaria digitata being the most rapidly hydrolysed substrate of those tested. K(m) values for GN I, II, and III against L. digitata laminarin were 0.1, 0.23 and 0.22 mg/ml respectively. D-Glucono-1,5-lactone does not inhibit any of the three GNs, some metals ions are mild inhibitors, and N-bromosuccinimide and KMnO4 are strong inhibitors. All three GNs acted in an exo-hydrolytic manner, determined by the release of alpha-glucose as the initial and major product of hydrolysis of (1-->3)-beta-D-glucans, and confirmed by viscometric analysis and the inability to cleave periodate-oxidized laminarin, and may be classified as (1-->3)-beta-D-glucan glucohydrolases (EC 3.2.1.58).

Excreted/secreted products of developing Taenia saginata metacestodes

Parasitology ◽  
1988 ◽  
Vol 97 (3) ◽  
pp. 477-487 ◽  
Author(s):  
G. W. P. Joshua ◽  
L. J. S. Harrison ◽  
M. M. H. Sewell
Keyword(s):  
Taenia Saginata ◽  
Cellular Infiltrate ◽  
Diagnostic Assays ◽  
Antigenic Characterization ◽  
Sds Page ◽  
Host Parasite Relationship ◽  
Parasite Relationship ◽  
Host Parasite

SummaryExcretions and secretions (ES) and somatic components of 4, 8, 12 and 16-week-old Taenia saginata metacestodes were biosynthetically radio-isotope labelled by incubating the larvae in the presence of [35S]methionine. Despite their small size, 4-week-old metacestodes produced as much isotope-labelled ES/parasite as older metacestodes, indicating a proportionately greater metabolic activity of the parasite at this age. In situ the 4-week-old metacestodes were surrounded by a marked granulomatous cellular infiltrate which had largely resolved around 8-week-old metacestodes. Examination of the isotope-labelled ES by SDS-PAGE revealed distinct age-specific components from 4- and 12-week-old metacestodes and other ES components which were produced by all the ages of metacestodes examined. In comparison the labelled somatic components were conserved. Antigenic characterization of the ES by immunoprecipitation against a panel of clinically defined bovine sera combined with SDS-PAGE analysis, identified some highly immunogenic parasite products and others which did not elicit an antibody response demonstrable by immunoprecipitation. These components are of interest in relation to the host/parasite relationship, to the construction of diagnostic assays for the detection of T. saginata cysticercosis, and to the immunity that cattle develop against this parasite.

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