Ultrastructural localisation of lipopolysaccharide-binding sites with peroxidase-conjugated lipopolysaccharides

Nygren, Håkan; Dahlén, Gunnar

doi:10.1016/0022-1759(79)90027-9

Cited by 10 publications

(4 citation statements)

References 17 publications

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“…Labeling can be achieved with radiochemicals by metabolic incorporation [13][14][15][16], a laborious method with the typical concerns involved in the manipulation of radioactive material, or by chemical conjugation. Fluorescein isothiocyanate (FITC) [7,9], horseradish peroxidase (HRP) [11,17], and biotin [8,18], among other compounds, have been extensively used to label LPS; yet its conjugation poses some difficulties with respect to yield because of its chemical inertness.…”

Section: Introductionmentioning

confidence: 99%

An efficient method for conjugation of a lipopolysaccharide from Salmonella enterica sv. Minnesota with probes bearing hydrazine or amino functional groups

Pallarola

Battaglini

2008

Analytical Biochemistry

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Section: Introductionmentioning

confidence: 99%

An efficient method for conjugation of a lipopolysaccharide from Salmonella enterica sv. Minnesota with probes bearing hydrazine or amino functional groups

Pallarola

Battaglini

2008

Analytical Biochemistry

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“…Actually, it has been found thai T and B eells bind LPS equally well, although only B cells are activated [11]. In a previous study [12] it was found that unlabelled LPS will compete with horseradish-peroxidase-iabelled LPS for binding sites on cell membranes, suggesting that both sttbstances bind to the same receptors. Jn this paper we demonstrate that LPS conjugated to horseradish peroxidase is selectively bound lo a proportion of lymphocytes from the LPS highresponding strain C3H/Tif but not to lymphocytes from the C3H/HeJ low-responder strain, as judged by electron microscopy.…”

mentioning

confidence: 99%

Bacterial Lipopolysaccharides Bind Selectively to Lymphocytes from Lipopolysaccharide High‐responder Mouse Strains

Nygren¹,

Dahlén²,

Möller³

1979

Scand J Immunol

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Three different concentrations of horseradish peroxidase-labelled lipopolysaccharide (LPS-HRP) were added in vitro to spleen cells from the LPS high-responder strain C3H/Tif and to cells from the low-responder strain C3H/HeJ. After being washed and fixed the cells were exposed to the substrate and prepared for electron microscopy. After addition of 7 and 0.7 microgram/ml of labelled LPS only lymphocytes from the high-responder strain were labelled. About 5-10% of the cells from C3H/Tif bound LPS, which is in accordance with the known frequency of B cells possessing the genetically determined LPS receptor. At the highest dose of labelled LPS (70 microgram/ml) a large proportion of lymphocytes from the low-responder strain also bound LPS. Erythrocytes from both strains bound LPS at all concentrations. It is concluded that LPS-HRP allows the detection at the cellular level of LPS binding to the genetically controlled membrane receptor for LPS.

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“…Localization of endotoxic LPS to the erythrocyte plasma membrane has been demonstrated by thin-section electron microscopy, using peroxidase-conjugated LPS (17). A putative lipoglycoprotein receptor for LPS has been isolated TD 2 -= _ g from human erythrocyte membranes (21 of phosphatidylcholine, phosphatidylethanolamine, or phosphatidylserine and can also de-Effect of polymyxin B on stimulation of crease the stability of lecithin-cholesterol binphoid cells by LPS (endotoxin).…”

Section: Discussionmentioning

confidence: 99%

Polymyxin B suppresses the endotoxin inhibition of concanavalin a-mediated erythrocyte agglutination

Warren¹

1982

Infect Immun

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The lectin agglutinability of human erythrocytes has been utilized to examine interactions of gram-negative endotoxin with mammalian cell plasma membranes. Erythrocytes treated in buffer with Escherichia coli 0127: B8 lipopolysaccharide (LPS) or Salmonella minnesota Re595 glycolipid for 1 h became resistant to agglutination by the lectin concanavalin A (ConA) in buffer free of LPS or glycolipid. Polymyxin B, a cationic cyclic lipopeptide which specifically binds to the lipid A toxophore, was tested for possible effects on the LPS and glycolipid inhibition of ConA erythrocyte agglutination. The presence of polymyxin B during the initial 1-h treatment with LPS or glycolipid blocked the ability of the endotoxins to render erythrocytes refractory to agglutination by ConA. Inhibition by polymyxin B was stoichiometric, and in repeated experiments, LPS was completely suppressed in the hemagglutination assay at a polymyxin B:LPS weight ratio of 1:4.1 (increasing polymyxin concentration, constant LPS concentration) and 1:5.1 (constant polymyxin concentration, increasing LPS concentration). These stoichiometry values are similar to values obtained for inhibition by polymyxin B of LPS lymphoid cell activation. It was concluded, therefore, that endotoxin inhibition of ConA erythrocyte agglutination reflects interactions of erythrocyte membranes with the lipid A region of endotoxin. In addition, the stoichiometry of polymyxin B inhibition suggests a similar extent of lipid Adependent LPS interaction with erythrocytes and lymphoid cells.

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Ultrastructural localisation of lipopolysaccharide-binding sites with peroxidase-conjugated lipopolysaccharides

Cited by 10 publications

References 17 publications

An efficient method for conjugation of a lipopolysaccharide from Salmonella enterica sv. Minnesota with probes bearing hydrazine or amino functional groups

An efficient method for conjugation of a lipopolysaccharide from Salmonella enterica sv. Minnesota with probes bearing hydrazine or amino functional groups

Bacterial Lipopolysaccharides Bind Selectively to Lymphocytes from Lipopolysaccharide High‐responder Mouse Strains

Polymyxin B suppresses the endotoxin inhibition of concanavalin a-mediated erythrocyte agglutination

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