A flow cytometry evaluation of anti-FVIII antibodies: Correlation with ELISA and Bethesda assay

Autores
Irigoyen, María Belén; Primiani, L.; Felippo, Marta Elena; Candela, M.; Perez Bianco, R.; De Bracco, M. M. DE E.; Galassi, Nora Virginia
Año de publicación
2011
Idioma
inglés
Tipo de recurso
artículo
Estado
versión publicada
Descripción
In this study, we describe a flow cytometry (FC) system for detecting antibodies to factor VIII (FVIII) and compare its results with those of enzyme-linked immunosorbent assay (ELISA) that detects both inhibitory (I-Ab) and non-inhibitory (NI-Ab) antibodies and the Nijmegen modification of the Bethesda method, detecting I-Ab. FC was set up in our laboratory. Recombinant FVIII (rFVIII) was coupled to microspheres (FVIII-m) and reacted with different plasma dilutions. Microspheres without rFVIII were used as control (control-m). Captured anti-FVIII antibodies were detected using anti-human IgG. Plasma samples from the following patients with severe haemophilia A (SHA) patients were evaluated: 17 P (patients without I-Ab, <0.5BUmL-1); 13 PI (patients with I-Ab, 1.1-8200BUmL-1). Of these 13, two PI were referred during immune tolerance induction (ITI), and plasmas from 12 healthy donors (HD) were evaluated. Semiquantitative results were given as an index (the highest mean fluorescence intensity ratio between FVIII-m and control-m multiplied by the inverse of the corresponding plasma dilution). Both plasma and serum were suitable for the test. FC agreed with the Bethesda method (r=0.8; P=0.0001). FC and ELISA had 80% of coincidence. Four of 17 patients (23.5%) had NI-Ab by FC, and two of them developed high levels of I-Ab later on. This test provides a useful alternative for measuring FVIII antibodies supplementing Bethesda assay. FC is fast and easy to perform. No more than 200μL of plasma or serum is required especially making it useful for paediatric patients. © 2010 Blackwell Publishing Ltd.
Fil: Irigoyen, María Belén. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Academia Nacional de Medicina de Buenos Aires; Argentina
Fil: Primiani, L.. Fundación Argentina de Hemofilia; Argentina
Fil: Felippo, Marta Elena. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Academia Nacional de Medicina de Buenos Aires; Argentina
Fil: Candela, M.. Academia Nacional de Medicina de Buenos Aires; Argentina
Fil: Perez Bianco, R.. Academia Nacional de Medicina de Buenos Aires; Argentina
Fil: De Bracco, M. M. DE E.. Academia Nacional de Medicina de Buenos Aires; Argentina
Fil: Galassi, Nora Virginia. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Academia Nacional de Medicina de Buenos Aires; Argentina
Materia
Flow Cytometry
Fviii Antibodies
Haemophilia A
Microspheres
Nivel de accesibilidad
acceso abierto
Condiciones de uso
https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
Repositorio
CONICET Digital (CONICET)
Institución
Consejo Nacional de Investigaciones Científicas y Técnicas
OAI Identificador
oai:ri.conicet.gov.ar:11336/52932
Acceder
id CONICETDig_e78478017524154844446274799da3d3
oai_identifier_str oai:ri.conicet.gov.ar:11336/52932
network_acronym_str CONICETDig
repository_id_str 3498
network_name_str CONICET Digital (CONICET)
spelling A flow cytometry evaluation of anti-FVIII antibodies: Correlation with ELISA and Bethesda assayIrigoyen, María BelénPrimiani, L.Felippo, Marta ElenaCandela, M.Perez Bianco, R.De Bracco, M. M. DE E.Galassi, Nora VirginiaFlow CytometryFviii AntibodiesHaemophilia AMicrosphereshttps://purl.org/becyt/ford/3.2https://purl.org/becyt/ford/3In this study, we describe a flow cytometry (FC) system for detecting antibodies to factor VIII (FVIII) and compare its results with those of enzyme-linked immunosorbent assay (ELISA) that detects both inhibitory (I-Ab) and non-inhibitory (NI-Ab) antibodies and the Nijmegen modification of the Bethesda method, detecting I-Ab. FC was set up in our laboratory. Recombinant FVIII (rFVIII) was coupled to microspheres (FVIII-m) and reacted with different plasma dilutions. Microspheres without rFVIII were used as control (control-m). Captured anti-FVIII antibodies were detected using anti-human IgG. Plasma samples from the following patients with severe haemophilia A (SHA) patients were evaluated: 17 P (patients without I-Ab, <0.5BUmL-1); 13 PI (patients with I-Ab, 1.1-8200BUmL-1). Of these 13, two PI were referred during immune tolerance induction (ITI), and plasmas from 12 healthy donors (HD) were evaluated. Semiquantitative results were given as an index (the highest mean fluorescence intensity ratio between FVIII-m and control-m multiplied by the inverse of the corresponding plasma dilution). Both plasma and serum were suitable for the test. FC agreed with the Bethesda method (r=0.8; P=0.0001). FC and ELISA had 80% of coincidence. Four of 17 patients (23.5%) had NI-Ab by FC, and two of them developed high levels of I-Ab later on. This test provides a useful alternative for measuring FVIII antibodies supplementing Bethesda assay. FC is fast and easy to perform. No more than 200μL of plasma or serum is required especially making it useful for paediatric patients. © 2010 Blackwell Publishing Ltd.Fil: Irigoyen, María Belén. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Academia Nacional de Medicina de Buenos Aires; ArgentinaFil: Primiani, L.. Fundación Argentina de Hemofilia; ArgentinaFil: Felippo, Marta Elena. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Academia Nacional de Medicina de Buenos Aires; ArgentinaFil: Candela, M.. Academia Nacional de Medicina de Buenos Aires; ArgentinaFil: Perez Bianco, R.. Academia Nacional de Medicina de Buenos Aires; ArgentinaFil: De Bracco, M. M. DE E.. Academia Nacional de Medicina de Buenos Aires; ArgentinaFil: Galassi, Nora Virginia. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Academia Nacional de Medicina de Buenos Aires; ArgentinaWiley Blackwell Publishing, Inc2011-03info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersionhttp://purl.org/coar/resource_type/c_6501info:ar-repo/semantics/articuloapplication/pdfapplication/pdfhttp://hdl.handle.net/11336/52932Irigoyen, María Belén; Primiani, L.; Felippo, Marta Elena; Candela, M.; Perez Bianco, R.; et al.; A flow cytometry evaluation of anti-FVIII antibodies: Correlation with ELISA and Bethesda assay; Wiley Blackwell Publishing, Inc; Haemophilia The Official Journal Of The World Federation Of Hemophilia; 17; 2; 3-2011; 267-2741351-8216CONICET DigitalCONICETenginfo:eu-repo/semantics/altIdentifier/doi/10.1111/j.1365-2516.2010.02406.xinfo:eu-repo/semantics/altIdentifier/url/https://onlinelibrary.wiley.com/doi/abs/10.1111/j.1365-2516.2010.02406.xinfo:eu-repo/semantics/openAccesshttps://creativecommons.org/licenses/by-nc-sa/2.5/ar/reponame:CONICET Digital (CONICET)instname:Consejo Nacional de Investigaciones Científicas y Técnicas2025-09-03T09:49:01Zoai:ri.conicet.gov.ar:11336/52932instacron:CONICETInstitucionalhttp://ri.conicet.gov.ar/Organismo científico-tecnológicoNo correspondehttp://ri.conicet.gov.ar/oai/requestdasensio@conicet.gov.ar; lcarlino@conicet.gov.arArgentinaNo correspondeNo correspondeNo correspondeopendoar:34982025-09-03 09:49:02.39CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicasfalse
dc.title.none.fl_str_mv A flow cytometry evaluation of anti-FVIII antibodies: Correlation with ELISA and Bethesda assay
title A flow cytometry evaluation of anti-FVIII antibodies: Correlation with ELISA and Bethesda assay
spellingShingle A flow cytometry evaluation of anti-FVIII antibodies: Correlation with ELISA and Bethesda assay
Irigoyen, María Belén
Flow Cytometry
Fviii Antibodies
Haemophilia A
Microspheres
title_short A flow cytometry evaluation of anti-FVIII antibodies: Correlation with ELISA and Bethesda assay
title_full A flow cytometry evaluation of anti-FVIII antibodies: Correlation with ELISA and Bethesda assay
title_fullStr A flow cytometry evaluation of anti-FVIII antibodies: Correlation with ELISA and Bethesda assay
title_full_unstemmed A flow cytometry evaluation of anti-FVIII antibodies: Correlation with ELISA and Bethesda assay
title_sort A flow cytometry evaluation of anti-FVIII antibodies: Correlation with ELISA and Bethesda assay
dc.creator.none.fl_str_mv Irigoyen, María Belén
Primiani, L.
Felippo, Marta Elena
Candela, M.
Perez Bianco, R.
De Bracco, M. M. DE E.
Galassi, Nora Virginia
author Irigoyen, María Belén
author_facet Irigoyen, María Belén
Primiani, L.
Felippo, Marta Elena
Candela, M.
Perez Bianco, R.
De Bracco, M. M. DE E.
Galassi, Nora Virginia
author_role author
author2 Primiani, L.
Felippo, Marta Elena
Candela, M.
Perez Bianco, R.
De Bracco, M. M. DE E.
Galassi, Nora Virginia
author2_role author
author
author
author
author
author
dc.subject.none.fl_str_mv Flow Cytometry
Fviii Antibodies
Haemophilia A
Microspheres
topic Flow Cytometry
Fviii Antibodies
Haemophilia A
Microspheres
purl_subject.fl_str_mv https://purl.org/becyt/ford/3.2
https://purl.org/becyt/ford/3
dc.description.none.fl_txt_mv In this study, we describe a flow cytometry (FC) system for detecting antibodies to factor VIII (FVIII) and compare its results with those of enzyme-linked immunosorbent assay (ELISA) that detects both inhibitory (I-Ab) and non-inhibitory (NI-Ab) antibodies and the Nijmegen modification of the Bethesda method, detecting I-Ab. FC was set up in our laboratory. Recombinant FVIII (rFVIII) was coupled to microspheres (FVIII-m) and reacted with different plasma dilutions. Microspheres without rFVIII were used as control (control-m). Captured anti-FVIII antibodies were detected using anti-human IgG. Plasma samples from the following patients with severe haemophilia A (SHA) patients were evaluated: 17 P (patients without I-Ab, <0.5BUmL-1); 13 PI (patients with I-Ab, 1.1-8200BUmL-1). Of these 13, two PI were referred during immune tolerance induction (ITI), and plasmas from 12 healthy donors (HD) were evaluated. Semiquantitative results were given as an index (the highest mean fluorescence intensity ratio between FVIII-m and control-m multiplied by the inverse of the corresponding plasma dilution). Both plasma and serum were suitable for the test. FC agreed with the Bethesda method (r=0.8; P=0.0001). FC and ELISA had 80% of coincidence. Four of 17 patients (23.5%) had NI-Ab by FC, and two of them developed high levels of I-Ab later on. This test provides a useful alternative for measuring FVIII antibodies supplementing Bethesda assay. FC is fast and easy to perform. No more than 200μL of plasma or serum is required especially making it useful for paediatric patients. © 2010 Blackwell Publishing Ltd.
Fil: Irigoyen, María Belén. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Academia Nacional de Medicina de Buenos Aires; Argentina
Fil: Primiani, L.. Fundación Argentina de Hemofilia; Argentina
Fil: Felippo, Marta Elena. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Academia Nacional de Medicina de Buenos Aires; Argentina
Fil: Candela, M.. Academia Nacional de Medicina de Buenos Aires; Argentina
Fil: Perez Bianco, R.. Academia Nacional de Medicina de Buenos Aires; Argentina
Fil: De Bracco, M. M. DE E.. Academia Nacional de Medicina de Buenos Aires; Argentina
Fil: Galassi, Nora Virginia. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina. Academia Nacional de Medicina de Buenos Aires; Argentina
description In this study, we describe a flow cytometry (FC) system for detecting antibodies to factor VIII (FVIII) and compare its results with those of enzyme-linked immunosorbent assay (ELISA) that detects both inhibitory (I-Ab) and non-inhibitory (NI-Ab) antibodies and the Nijmegen modification of the Bethesda method, detecting I-Ab. FC was set up in our laboratory. Recombinant FVIII (rFVIII) was coupled to microspheres (FVIII-m) and reacted with different plasma dilutions. Microspheres without rFVIII were used as control (control-m). Captured anti-FVIII antibodies were detected using anti-human IgG. Plasma samples from the following patients with severe haemophilia A (SHA) patients were evaluated: 17 P (patients without I-Ab, <0.5BUmL-1); 13 PI (patients with I-Ab, 1.1-8200BUmL-1). Of these 13, two PI were referred during immune tolerance induction (ITI), and plasmas from 12 healthy donors (HD) were evaluated. Semiquantitative results were given as an index (the highest mean fluorescence intensity ratio between FVIII-m and control-m multiplied by the inverse of the corresponding plasma dilution). Both plasma and serum were suitable for the test. FC agreed with the Bethesda method (r=0.8; P=0.0001). FC and ELISA had 80% of coincidence. Four of 17 patients (23.5%) had NI-Ab by FC, and two of them developed high levels of I-Ab later on. This test provides a useful alternative for measuring FVIII antibodies supplementing Bethesda assay. FC is fast and easy to perform. No more than 200μL of plasma or serum is required especially making it useful for paediatric patients. © 2010 Blackwell Publishing Ltd.
publishDate 2011
dc.date.none.fl_str_mv 2011-03
dc.type.none.fl_str_mv info:eu-repo/semantics/article
info:eu-repo/semantics/publishedVersion
http://purl.org/coar/resource_type/c_6501
info:ar-repo/semantics/articulo
format article
status_str publishedVersion
dc.identifier.none.fl_str_mv http://hdl.handle.net/11336/52932
Irigoyen, María Belén; Primiani, L.; Felippo, Marta Elena; Candela, M.; Perez Bianco, R.; et al.; A flow cytometry evaluation of anti-FVIII antibodies: Correlation with ELISA and Bethesda assay; Wiley Blackwell Publishing, Inc; Haemophilia The Official Journal Of The World Federation Of Hemophilia; 17; 2; 3-2011; 267-274
1351-8216
CONICET Digital
CONICET
url http://hdl.handle.net/11336/52932
identifier_str_mv Irigoyen, María Belén; Primiani, L.; Felippo, Marta Elena; Candela, M.; Perez Bianco, R.; et al.; A flow cytometry evaluation of anti-FVIII antibodies: Correlation with ELISA and Bethesda assay; Wiley Blackwell Publishing, Inc; Haemophilia The Official Journal Of The World Federation Of Hemophilia; 17; 2; 3-2011; 267-274
1351-8216
CONICET Digital
CONICET
dc.language.none.fl_str_mv eng
language eng
dc.relation.none.fl_str_mv info:eu-repo/semantics/altIdentifier/doi/10.1111/j.1365-2516.2010.02406.x
info:eu-repo/semantics/altIdentifier/url/https://onlinelibrary.wiley.com/doi/abs/10.1111/j.1365-2516.2010.02406.x
dc.rights.none.fl_str_mv info:eu-repo/semantics/openAccess
https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
eu_rights_str_mv openAccess
rights_invalid_str_mv https://creativecommons.org/licenses/by-nc-sa/2.5/ar/
dc.format.none.fl_str_mv application/pdf
application/pdf
dc.publisher.none.fl_str_mv Wiley Blackwell Publishing, Inc
publisher.none.fl_str_mv Wiley Blackwell Publishing, Inc
dc.source.none.fl_str_mv reponame:CONICET Digital (CONICET)
instname:Consejo Nacional de Investigaciones Científicas y Técnicas
reponame_str CONICET Digital (CONICET)
collection CONICET Digital (CONICET)
instname_str Consejo Nacional de Investigaciones Científicas y Técnicas
repository.name.fl_str_mv CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicas
repository.mail.fl_str_mv dasensio@conicet.gov.ar; lcarlino@conicet.gov.ar
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