Development and evaluation of a novel VHH-based immunocapture assay for high-sensitivity detection of Shiga toxin type 2 (Stx2) in stool samples

Autores
Melli, Luciano Jorge; Zylberman, Vanesa; Hiriart, Yanina; Lauche, Constanza E.; Baschkier, Ariela; Pardo, Romina; Miliwebsky, Elizabeth; Chinen, Isabel; Rivas, Marta; Goldbaum, Fernando A.; Ugalde, Juan Esteban; Comerci, Diego José; Ciocchini, Andres Eduardo
Año de publicación
2020
Idioma
inglés
Tipo de recurso
artículo
Estado
versión publicada
Descripción
Shiga toxin (Stx)-producing Escherichia coli (STEC) is the main cause of postdiarrheal hemolytic-uremic syndrome (HUS), a life-threatening clinical complication characterized by hemolytic anemia, thrombocytopenia, and acute renal failure that mainly affects children. A relevant feature of STEC strains is the production of Stx, and all of them express Stx1 and/or Stx2 regardless of the strain serotype. Therefore, Stx detection assays are considered the most suitable methods for the early detection of STEC infections. Single-domain antibodies from camelids (VHHs) exhibit several advantages in comparison with conventional antibodies, making them promising tools for diagnosis. In this work, we have exploited VHH technology for the development of an immunocapture assay for Stx2 detection. Thirteen anti- Stx2 VHHs previously obtained from a variable-domain repertoire library were selected and evaluated in 130 capture-detection pair combinations for Stx detection. Based on this analysis, two VHHs were selected and a double VHH-based biotinstreptavidin capture enzyme-linked immunosorbent assay (ELISA) with spectrophotometric detection was developed and optimized for Stx2 detection. This assay showed an excellent analytical and clinical sensitivity in both STEC culture supernatants and stool samples even higher than the sensitivity of a commercial ELISA. Furthermore, based on the analysis of stool samples, the VHH-based ELISA showed high correlation with stx2 detection by PCR and a commercial rapid membrane-based immunoassay. The intrinsic properties of VHHs (high target affinity and specificity, stability, and ease of expression at high yields in recombinant bacteria) and their optimal performance for Stx detection make them attractive tools for the diagnosis of HUS related to STEC (STEC-HUS).
Fil: Melli, Luciano Jorge. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Biotecnológicas; Argentina
Fil: Zylberman, Vanesa. Inmunova; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
Fil: Hiriart, Yanina. Inmunova; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
Fil: Lauche, Constanza E.. Inmunova; Argentina
Fil: Baschkier, Ariela. Dirección Nacional de Institutos de Investigación. Administración Nacional de Laboratorios e Institutos de Salud. Instituto Nacional de Enfermedades Infecciosas; Argentina
Fil: Pardo, Romina. Inmunova; Argentina
Fil: Miliwebsky, Elizabeth. Dirección Nacional de Institutos de Investigación. Administración Nacional de Laboratorios e Institutos de Salud. Instituto Nacional de Enfermedades Infecciosas; Argentina
Fil: Chinen, Isabel. Dirección Nacional de Institutos de Investigación. Administración Nacional de Laboratorios e Institutos de Salud. Instituto Nacional de Enfermedades Infecciosas; Argentina
Fil: Rivas, Marta. Dirección Nacional de Institutos de Investigación. Administración Nacional de Laboratorios e Institutos de Salud. Instituto Nacional de Enfermedades Infecciosas; Argentina
Fil: Goldbaum, Fernando A.. Inmunova; Argentina
Fil: Ugalde, Juan Esteban. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Biotecnológicas; Argentina
Fil: Comerci, Diego José. Inmunova; Argentina. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Biotecnológicas; Argentina
Fil: Ciocchini, Andres Eduardo. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Biotecnológicas; Argentina
Materia
ELISA
HUS
NANOBODIES
SHIGA TOXIN
STEC
STX2
VHH
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:11336/140320
Acceder
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oai_identifier_str oai:ri.conicet.gov.ar:11336/140320
network_acronym_str CONICETDig
repository_id_str 3498
network_name_str CONICET Digital (CONICET)
spelling Development and evaluation of a novel VHH-based immunocapture assay for high-sensitivity detection of Shiga toxin type 2 (Stx2) in stool samplesMelli, Luciano JorgeZylberman, VanesaHiriart, YaninaLauche, Constanza E.Baschkier, ArielaPardo, RominaMiliwebsky, ElizabethChinen, IsabelRivas, MartaGoldbaum, Fernando A.Ugalde, Juan EstebanComerci, Diego JoséCiocchini, Andres EduardoELISAHUSNANOBODIESSHIGA TOXINSTECSTX2VHHhttps://purl.org/becyt/ford/1.6https://purl.org/becyt/ford/1Shiga toxin (Stx)-producing Escherichia coli (STEC) is the main cause of postdiarrheal hemolytic-uremic syndrome (HUS), a life-threatening clinical complication characterized by hemolytic anemia, thrombocytopenia, and acute renal failure that mainly affects children. A relevant feature of STEC strains is the production of Stx, and all of them express Stx1 and/or Stx2 regardless of the strain serotype. Therefore, Stx detection assays are considered the most suitable methods for the early detection of STEC infections. Single-domain antibodies from camelids (VHHs) exhibit several advantages in comparison with conventional antibodies, making them promising tools for diagnosis. In this work, we have exploited VHH technology for the development of an immunocapture assay for Stx2 detection. Thirteen anti- Stx2 VHHs previously obtained from a variable-domain repertoire library were selected and evaluated in 130 capture-detection pair combinations for Stx detection. Based on this analysis, two VHHs were selected and a double VHH-based biotinstreptavidin capture enzyme-linked immunosorbent assay (ELISA) with spectrophotometric detection was developed and optimized for Stx2 detection. This assay showed an excellent analytical and clinical sensitivity in both STEC culture supernatants and stool samples even higher than the sensitivity of a commercial ELISA. Furthermore, based on the analysis of stool samples, the VHH-based ELISA showed high correlation with stx2 detection by PCR and a commercial rapid membrane-based immunoassay. The intrinsic properties of VHHs (high target affinity and specificity, stability, and ease of expression at high yields in recombinant bacteria) and their optimal performance for Stx detection make them attractive tools for the diagnosis of HUS related to STEC (STEC-HUS).Fil: Melli, Luciano Jorge. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Biotecnológicas; ArgentinaFil: Zylberman, Vanesa. Inmunova; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Hiriart, Yanina. Inmunova; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: Lauche, Constanza E.. Inmunova; ArgentinaFil: Baschkier, Ariela. Dirección Nacional de Institutos de Investigación. Administración Nacional de Laboratorios e Institutos de Salud. Instituto Nacional de Enfermedades Infecciosas; ArgentinaFil: Pardo, Romina. Inmunova; ArgentinaFil: Miliwebsky, Elizabeth. Dirección Nacional de Institutos de Investigación. Administración Nacional de Laboratorios e Institutos de Salud. Instituto Nacional de Enfermedades Infecciosas; ArgentinaFil: Chinen, Isabel. Dirección Nacional de Institutos de Investigación. Administración Nacional de Laboratorios e Institutos de Salud. Instituto Nacional de Enfermedades Infecciosas; ArgentinaFil: Rivas, Marta. Dirección Nacional de Institutos de Investigación. Administración Nacional de Laboratorios e Institutos de Salud. Instituto Nacional de Enfermedades Infecciosas; ArgentinaFil: Goldbaum, Fernando A.. Inmunova; ArgentinaFil: Ugalde, Juan Esteban. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Biotecnológicas; ArgentinaFil: Comerci, Diego José. Inmunova; Argentina. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Biotecnológicas; ArgentinaFil: Ciocchini, Andres Eduardo. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Biotecnológicas; ArgentinaAmerican Society for Microbiology2020-02info:eu-repo/semantics/articleinfo:eu-repo/semantics/publishedVersionhttp://purl.org/coar/resource_type/c_6501info:ar-repo/semantics/articuloapplication/pdfapplication/pdfhttp://hdl.handle.net/11336/140320Melli, Luciano Jorge; Zylberman, Vanesa; Hiriart, Yanina; Lauche, Constanza E.; Baschkier, Ariela; et al.; Development and evaluation of a novel VHH-based immunocapture assay for high-sensitivity detection of Shiga toxin type 2 (Stx2) in stool samples; American Society for Microbiology; Journal of Clinical Microbiology; 58; 3; 2-2020; 1-150095-1137CONICET DigitalCONICETenginfo:eu-repo/semantics/altIdentifier/url/https://jcm.asm.org/content/58/3/e01566-19info:eu-repo/semantics/altIdentifier/doi/10.1128/JCM.01566-19info:eu-repo/semantics/openAccesshttps://creativecommons.org/licenses/by-nc-sa/2.5/ar/reponame:CONICET Digital (CONICET)instname:Consejo Nacional de Investigaciones Científicas y Técnicas2025-09-03T09:56:31Zoai:ri.conicet.gov.ar:11336/140320instacron: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 09:56:31.536CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicasfalse
dc.title.none.fl_str_mv Development and evaluation of a novel VHH-based immunocapture assay for high-sensitivity detection of Shiga toxin type 2 (Stx2) in stool samples
title Development and evaluation of a novel VHH-based immunocapture assay for high-sensitivity detection of Shiga toxin type 2 (Stx2) in stool samples
spellingShingle Development and evaluation of a novel VHH-based immunocapture assay for high-sensitivity detection of Shiga toxin type 2 (Stx2) in stool samples
Melli, Luciano Jorge
ELISA
HUS
NANOBODIES
SHIGA TOXIN
STEC
STX2
VHH
title_short Development and evaluation of a novel VHH-based immunocapture assay for high-sensitivity detection of Shiga toxin type 2 (Stx2) in stool samples
title_full Development and evaluation of a novel VHH-based immunocapture assay for high-sensitivity detection of Shiga toxin type 2 (Stx2) in stool samples
title_fullStr Development and evaluation of a novel VHH-based immunocapture assay for high-sensitivity detection of Shiga toxin type 2 (Stx2) in stool samples
title_full_unstemmed Development and evaluation of a novel VHH-based immunocapture assay for high-sensitivity detection of Shiga toxin type 2 (Stx2) in stool samples
title_sort Development and evaluation of a novel VHH-based immunocapture assay for high-sensitivity detection of Shiga toxin type 2 (Stx2) in stool samples
dc.creator.none.fl_str_mv Melli, Luciano Jorge
Zylberman, Vanesa
Hiriart, Yanina
Lauche, Constanza E.
Baschkier, Ariela
Pardo, Romina
Miliwebsky, Elizabeth
Chinen, Isabel
Rivas, Marta
Goldbaum, Fernando A.
Ugalde, Juan Esteban
Comerci, Diego José
Ciocchini, Andres Eduardo
author Melli, Luciano Jorge
author_facet Melli, Luciano Jorge
Zylberman, Vanesa
Hiriart, Yanina
Lauche, Constanza E.
Baschkier, Ariela
Pardo, Romina
Miliwebsky, Elizabeth
Chinen, Isabel
Rivas, Marta
Goldbaum, Fernando A.
Ugalde, Juan Esteban
Comerci, Diego José
Ciocchini, Andres Eduardo
author_role author
author2 Zylberman, Vanesa
Hiriart, Yanina
Lauche, Constanza E.
Baschkier, Ariela
Pardo, Romina
Miliwebsky, Elizabeth
Chinen, Isabel
Rivas, Marta
Goldbaum, Fernando A.
Ugalde, Juan Esteban
Comerci, Diego José
Ciocchini, Andres Eduardo
author2_role author
author
author
author
author
author
author
author
author
author
author
author
dc.subject.none.fl_str_mv ELISA
HUS
NANOBODIES
SHIGA TOXIN
STEC
STX2
VHH
topic ELISA
HUS
NANOBODIES
SHIGA TOXIN
STEC
STX2
VHH
purl_subject.fl_str_mv https://purl.org/becyt/ford/1.6
https://purl.org/becyt/ford/1
dc.description.none.fl_txt_mv Shiga toxin (Stx)-producing Escherichia coli (STEC) is the main cause of postdiarrheal hemolytic-uremic syndrome (HUS), a life-threatening clinical complication characterized by hemolytic anemia, thrombocytopenia, and acute renal failure that mainly affects children. A relevant feature of STEC strains is the production of Stx, and all of them express Stx1 and/or Stx2 regardless of the strain serotype. Therefore, Stx detection assays are considered the most suitable methods for the early detection of STEC infections. Single-domain antibodies from camelids (VHHs) exhibit several advantages in comparison with conventional antibodies, making them promising tools for diagnosis. In this work, we have exploited VHH technology for the development of an immunocapture assay for Stx2 detection. Thirteen anti- Stx2 VHHs previously obtained from a variable-domain repertoire library were selected and evaluated in 130 capture-detection pair combinations for Stx detection. Based on this analysis, two VHHs were selected and a double VHH-based biotinstreptavidin capture enzyme-linked immunosorbent assay (ELISA) with spectrophotometric detection was developed and optimized for Stx2 detection. This assay showed an excellent analytical and clinical sensitivity in both STEC culture supernatants and stool samples even higher than the sensitivity of a commercial ELISA. Furthermore, based on the analysis of stool samples, the VHH-based ELISA showed high correlation with stx2 detection by PCR and a commercial rapid membrane-based immunoassay. The intrinsic properties of VHHs (high target affinity and specificity, stability, and ease of expression at high yields in recombinant bacteria) and their optimal performance for Stx detection make them attractive tools for the diagnosis of HUS related to STEC (STEC-HUS).
Fil: Melli, Luciano Jorge. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Biotecnológicas; Argentina
Fil: Zylberman, Vanesa. Inmunova; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
Fil: Hiriart, Yanina. Inmunova; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; Argentina
Fil: Lauche, Constanza E.. Inmunova; Argentina
Fil: Baschkier, Ariela. Dirección Nacional de Institutos de Investigación. Administración Nacional de Laboratorios e Institutos de Salud. Instituto Nacional de Enfermedades Infecciosas; Argentina
Fil: Pardo, Romina. Inmunova; Argentina
Fil: Miliwebsky, Elizabeth. Dirección Nacional de Institutos de Investigación. Administración Nacional de Laboratorios e Institutos de Salud. Instituto Nacional de Enfermedades Infecciosas; Argentina
Fil: Chinen, Isabel. Dirección Nacional de Institutos de Investigación. Administración Nacional de Laboratorios e Institutos de Salud. Instituto Nacional de Enfermedades Infecciosas; Argentina
Fil: Rivas, Marta. Dirección Nacional de Institutos de Investigación. Administración Nacional de Laboratorios e Institutos de Salud. Instituto Nacional de Enfermedades Infecciosas; Argentina
Fil: Goldbaum, Fernando A.. Inmunova; Argentina
Fil: Ugalde, Juan Esteban. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Biotecnológicas; Argentina
Fil: Comerci, Diego José. Inmunova; Argentina. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Biotecnológicas; Argentina
Fil: Ciocchini, Andres Eduardo. Universidad Nacional de San Martín. Instituto de Investigaciones Biotecnológicas. - Consejo Nacional de Investigaciones Científicas y Técnicas. Oficina de Coordinación Administrativa Parque Centenario. Instituto de Investigaciones Biotecnológicas; Argentina
description Shiga toxin (Stx)-producing Escherichia coli (STEC) is the main cause of postdiarrheal hemolytic-uremic syndrome (HUS), a life-threatening clinical complication characterized by hemolytic anemia, thrombocytopenia, and acute renal failure that mainly affects children. A relevant feature of STEC strains is the production of Stx, and all of them express Stx1 and/or Stx2 regardless of the strain serotype. Therefore, Stx detection assays are considered the most suitable methods for the early detection of STEC infections. Single-domain antibodies from camelids (VHHs) exhibit several advantages in comparison with conventional antibodies, making them promising tools for diagnosis. In this work, we have exploited VHH technology for the development of an immunocapture assay for Stx2 detection. Thirteen anti- Stx2 VHHs previously obtained from a variable-domain repertoire library were selected and evaluated in 130 capture-detection pair combinations for Stx detection. Based on this analysis, two VHHs were selected and a double VHH-based biotinstreptavidin capture enzyme-linked immunosorbent assay (ELISA) with spectrophotometric detection was developed and optimized for Stx2 detection. This assay showed an excellent analytical and clinical sensitivity in both STEC culture supernatants and stool samples even higher than the sensitivity of a commercial ELISA. Furthermore, based on the analysis of stool samples, the VHH-based ELISA showed high correlation with stx2 detection by PCR and a commercial rapid membrane-based immunoassay. The intrinsic properties of VHHs (high target affinity and specificity, stability, and ease of expression at high yields in recombinant bacteria) and their optimal performance for Stx detection make them attractive tools for the diagnosis of HUS related to STEC (STEC-HUS).
publishDate 2020
dc.date.none.fl_str_mv 2020-02
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/11336/140320
Melli, Luciano Jorge; Zylberman, Vanesa; Hiriart, Yanina; Lauche, Constanza E.; Baschkier, Ariela; et al.; Development and evaluation of a novel VHH-based immunocapture assay for high-sensitivity detection of Shiga toxin type 2 (Stx2) in stool samples; American Society for Microbiology; Journal of Clinical Microbiology; 58; 3; 2-2020; 1-15
0095-1137
CONICET Digital
CONICET
url http://hdl.handle.net/11336/140320
identifier_str_mv Melli, Luciano Jorge; Zylberman, Vanesa; Hiriart, Yanina; Lauche, Constanza E.; Baschkier, Ariela; et al.; Development and evaluation of a novel VHH-based immunocapture assay for high-sensitivity detection of Shiga toxin type 2 (Stx2) in stool samples; American Society for Microbiology; Journal of Clinical Microbiology; 58; 3; 2-2020; 1-15
0095-1137
CONICET Digital
CONICET
dc.language.none.fl_str_mv eng
language eng
dc.relation.none.fl_str_mv info:eu-repo/semantics/altIdentifier/url/https://jcm.asm.org/content/58/3/e01566-19
info:eu-repo/semantics/altIdentifier/doi/10.1128/JCM.01566-19
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 American Society for Microbiology
publisher.none.fl_str_mv American Society for Microbiology
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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score 13.13397

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