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Article: Biotinylated lipopolysaccharide binds to endotoxin receptor in endothelial and monocytic cells

TitleBiotinylated lipopolysaccharide binds to endotoxin receptor in endothelial and monocytic cells
Authors
Issue Date1995
PublisherAcademic Press. The Journal's web site is located at http://www.elsevier.com/locate/yabio
Citation
Analytical Biochemistry, 1995, v. 232 n. 2, p. 217-224 How to Cite?
AbstractEndotoxin or lipopolysaccharide (LPS), a major cell surface component of gram-negative bacteria, which could bind to different cell types when released into the bloodstream, plays a central role in the pathogenesis of septic shock syndromes. We have established a biotinylation procedure for labeling purified LPS molecules from Salmonella minnesota R595 and Escherichia coli J5 bacteria. The biotin group was conjugated to the bacterial LPS either by chemical oxidation of the LPS carbohydrate moiety (inner core region), followed by reduction with biotin-LC-hydrazide (biotinamido hexanoyl hydrazide), or by photoactivatable cross-linking with biotin-LC-ASA [1-(4-azidosalicylamido-)-6(biotinamido)-hexane], which was randomly attached to the carbohydrate and fatty acid (lipid A) groups of the LPS. Both labeled products retained biological activity (or endotoxicity) as evidenced by coagulation of the Limulus amoebocyte lysate. To determine its ability to bind avidin/streptavidin which in turn could be conjugated with enzymatic and fluorescent probes, the biotinylated LPS was used in enzyme immunoassay, Western blot, and flow cytometry. These assays were also used to analyze the binding of LPS ligand to its counterreceptor(s) on whole cell surface, membrane fragments, and in detergent lysates from human endothelial and monocytic cells. The described biotinylated LPS probes can be applied in a wide variety of techniques in receptor biochemistry, immunohistochemistry, and molecular cell biology.
Persistent Identifierhttp://hdl.handle.net/10722/172719
ISSN
2015 Impact Factor: 2.243
2015 SCImago Journal Rankings: 0.725
ISI Accession Number ID

 

DC FieldValueLanguage
dc.contributor.authorLuk, JMen_US
dc.contributor.authorKumar, Aen_US
dc.contributor.authorTsang, Ren_US
dc.contributor.authorStaunton, Den_US
dc.date.accessioned2012-10-30T06:24:28Z-
dc.date.available2012-10-30T06:24:28Z-
dc.date.issued1995en_US
dc.identifier.citationAnalytical Biochemistry, 1995, v. 232 n. 2, p. 217-224en_US
dc.identifier.issn0003-2697en_US
dc.identifier.urihttp://hdl.handle.net/10722/172719-
dc.description.abstractEndotoxin or lipopolysaccharide (LPS), a major cell surface component of gram-negative bacteria, which could bind to different cell types when released into the bloodstream, plays a central role in the pathogenesis of septic shock syndromes. We have established a biotinylation procedure for labeling purified LPS molecules from Salmonella minnesota R595 and Escherichia coli J5 bacteria. The biotin group was conjugated to the bacterial LPS either by chemical oxidation of the LPS carbohydrate moiety (inner core region), followed by reduction with biotin-LC-hydrazide (biotinamido hexanoyl hydrazide), or by photoactivatable cross-linking with biotin-LC-ASA [1-(4-azidosalicylamido-)-6(biotinamido)-hexane], which was randomly attached to the carbohydrate and fatty acid (lipid A) groups of the LPS. Both labeled products retained biological activity (or endotoxicity) as evidenced by coagulation of the Limulus amoebocyte lysate. To determine its ability to bind avidin/streptavidin which in turn could be conjugated with enzymatic and fluorescent probes, the biotinylated LPS was used in enzyme immunoassay, Western blot, and flow cytometry. These assays were also used to analyze the binding of LPS ligand to its counterreceptor(s) on whole cell surface, membrane fragments, and in detergent lysates from human endothelial and monocytic cells. The described biotinylated LPS probes can be applied in a wide variety of techniques in receptor biochemistry, immunohistochemistry, and molecular cell biology.en_US
dc.languageengen_US
dc.publisherAcademic Press. The Journal's web site is located at http://www.elsevier.com/locate/yabioen_US
dc.relation.ispartofAnalytical Biochemistryen_US
dc.subject.meshAvidinen_US
dc.subject.meshAzidesen_US
dc.subject.meshBacterial Toxins - Metabolismen_US
dc.subject.meshBiotin - Analogs & Derivativesen_US
dc.subject.meshBlotting, Westernen_US
dc.subject.meshBordetella Pertussis - Chemistryen_US
dc.subject.meshCells, Cultureden_US
dc.subject.meshCross-Linking Reagentsen_US
dc.subject.meshDetergents - Pharmacologyen_US
dc.subject.meshElectrophoresis, Polyacrylamide Gelen_US
dc.subject.meshEndothelium, Vascular - Metabolismen_US
dc.subject.meshEndotoxins - Metabolismen_US
dc.subject.meshFlow Cytometryen_US
dc.subject.meshHl-60 Cells - Metabolismen_US
dc.subject.meshHumansen_US
dc.subject.meshImmunoenzyme Techniquesen_US
dc.subject.meshLimulus Testen_US
dc.subject.meshLipopolysaccharides - Metabolismen_US
dc.subject.meshMonocytes - Metabolismen_US
dc.subject.meshOxidation-Reductionen_US
dc.subject.meshPhotochemistryen_US
dc.subject.meshProtein Bindingen_US
dc.subject.meshReceptors, Immunologic - Metabolismen_US
dc.subject.meshRhodobacter Sphaeroides - Chemistryen_US
dc.subject.meshTumor Cells, Cultureden_US
dc.titleBiotinylated lipopolysaccharide binds to endotoxin receptor in endothelial and monocytic cellsen_US
dc.typeArticleen_US
dc.identifier.emailLuk, JM: jmluk@hkucc.hku.hken_US
dc.identifier.authorityLuk, JM=rp00349en_US
dc.description.naturelink_to_subscribed_fulltexten_US
dc.identifier.doi10.1006/abio.1995.0010en_US
dc.identifier.pmid8747478-
dc.identifier.scopuseid_2-s2.0-0029586188en_US
dc.identifier.volume232en_US
dc.identifier.issue2en_US
dc.identifier.spage217en_US
dc.identifier.epage224en_US
dc.identifier.isiWOS:A1995TM18900010-
dc.publisher.placeUnited Statesen_US
dc.identifier.scopusauthoridLuk, JM=7006777791en_US
dc.identifier.scopusauthoridKumar, A=7408039645en_US
dc.identifier.scopusauthoridTsang, R=7102940066en_US
dc.identifier.scopusauthoridStaunton, D=7005167471en_US

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