Simultaneous isolation of platelet factor 4 and glycoprotein IIb-IIIa complex from rabbit platelets, and characterization of specific chicken antibodies to assay them

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dc.contributor.authorSantoro, M. L.
dc.contributor.authorBarbaro, K. C.
dc.contributor.authorRocha, TRF da
dc.contributor.authorTorquato, RJS
dc.contributor.authorHirata, I. Y.
dc.contributor.authorSano-Martins, I. S.
dc.contributor.institutionInst Butantan
dc.contributor.institutionPro Sangue Fdn
dc.contributor.institutionUniversidade Federal de São Paulo (UNIFESP)
dc.date.accessioned2016-01-24T12:34:12Z
dc.date.available2016-01-24T12:34:12Z
dc.date.issued2004-01-01
dc.description.abstractRabbits are frequently used as models for studying coagulation and platelet disorders. However, few reports on literature have dealt with the purification and characterization of rabbit platelet proteins. Herein a protocol for the simultaneous purification of rabbit platelet factor 4 (PF4) and platelet glycoprotein IIb-IIIa (GPIIb-IIIa, integrin alpha(IIb)beta(3)) is described. Specific antibodies were raised in laying chicken, which were used for assaying PF4 by ELISA, and GPIIb-IIIa by direct immunofluorescence and flow cytometry. Furthermore, the binding of monoclonal antibodies specific for GPIIb-IIIa complex (P2), ligand-induced binding site of GPIIIa (LIBS1) and rabbit P-selectin (12A7), as well as of polyclonal IgY specific for rabbit GPIIb-IIIa, was compared in quiescent and thrombin-activated platelets. Polyclonal anti-rabbit PF4 IgY was a specific and sensitive probe that could be used for assaying PF4 in plasma samples. GPIIb-IIIa expression was increased in thrombin-activated platelets, as evaluated by flow cytometric analysis using P2 and polyclonal antibodies raised in chickens. Rabbit GPIIb-IIIa also exhibited a conformational modification that caused the appearance of ligand-induced binding sites. Increased P-selectin expression, used as a positive control, was also noticeable in thrombin-activated platelets. These data evidence that antibodies raised in laying chickens specific to rabbit PF4 and GPIIb-IIIa, as well as certain monoclonal antibodies specific for human GPIIb-IIIa, may be used for investigating rabbit platelet physiology. (C) 2003 Elsevier B.V. All rights reserved.en
dc.description.affiliationInst Butantan, Lab Pathophysiol, BR-05503900 São Paulo, SP, Brazil
dc.description.affiliationInst Butantan, Immunopathol Lab, BR-05503900 São Paulo, SP, Brazil
dc.description.affiliationPro Sangue Fdn, Hemoctr, Lab Thrombo Hemorrhag Dis, São Paulo, Brazil
dc.description.affiliationUniversidade Federal de São Paulo, Dept Biochem, São Paulo, SP, Brazil
dc.description.affiliationUniversidade Federal de São Paulo, Dept Biophys, São Paulo, SP, Brazil
dc.description.affiliationUnifespUniversidade Federal de São Paulo, Dept Biochem, São Paulo, SP, Brazil
dc.description.affiliationUnifespUniversidade Federal de São Paulo, Dept Biophys, São Paulo, SP, Brazil
dc.description.sourceWeb of Science
dc.format.extent55-72
dc.identifierhttp://dx.doi.org/10.1016/j.jim.2003.10.005
dc.identifier.citationJournal of Immunological Methods. Amsterdam: Elsevier B.V., v. 284, n. 1-2, p. 55-72, 2004.
dc.identifier.doi10.1016/j.jim.2003.10.005
dc.identifier.issn0022-1759
dc.identifier.urihttp://repositorio.unifesp.br/handle/11600/27541
dc.identifier.wosWOS:000188598400007
dc.language.isoeng
dc.publisherElsevier B.V.
dc.relation.ispartofJournal of Immunological Methods
dc.rightsinfo:eu-repo/semantics/restrictedAccess
dc.rights.licensehttp://www.elsevier.com/about/open-access/open-access-policies/article-posting-policy
dc.subjectchicken antibodiesen
dc.subjectplatelet factor 4en
dc.subjectglycoprotein IIb-IIIaen
dc.subjectintegrin alpha(IIb)beta(3)en
dc.subjectCD41en
dc.subjectCD61en
dc.subjectflow cytometryen
dc.subjectimmunoblottingen
dc.subjectIgYen
dc.subjectimmunofluorescenceen
dc.subjectrabbiten
dc.subjectthrombinen
dc.subjectP2en
dc.subject12A7en
dc.subjectLIBS1en
dc.subjectP-selectinen
dc.titleSimultaneous isolation of platelet factor 4 and glycoprotein IIb-IIIa complex from rabbit platelets, and characterization of specific chicken antibodies to assay themen
dc.typeinfo:eu-repo/semantics/article
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