Evaluation of an experimental botropic antivenom produced with venom blocked with Na2-EDTA

Autores
López, Gisela Lumila; Leiva, Laura Cristina Ana; Fusco, Luciano Sebastian
Año de publicación
2023
Idioma
inglés
Tipo de recurso
documento de conferencia
Estado
versión publicada
Descripción
Snake envenoming is a serious medical problem in tropical developing countries and antivenoms are the only effective therapy. Antisera are produced by immunizing horses with snake venom and adjuvant, usually Freund´s adjuvant. Botropic venoms contain metalloproteinases (SVMPs), responsible for the local effects of envenoming, such as hemorrhage, edema and myotoxicity as well as systemic bleeding. SVMPs represent around 34,2 % of the protein composition from B. diporus venom and causing lesions during the immunization of animals. In search of new processes, focused on animal welfare for the production immunobiologicals, and taking into account that the toxic action of SVMPs is inhibited by Na2-EDTA, in the present work, the immune response of animals inoculated with B.diporus venom blocked with Na2-EDTA was evaluated. For this proposal, the B. diporus venom (1,9 mg/mL) was blocked by Na2-EDTA (200 mM, B.dV/ Na2-EDTA) and used as antigen. Previously to the inoculation, the excess of chelate was removed by molecular exclusion chromatography (Sephadex G-25). Likewise, the venom without inhibitor (B.dV) received the same treatment and in both cases, the effective neutralization of SVMPs using azocasein as substrate was determined. Group of 5 CF-1 mice were immunized subcutaneously on days 0, 15 and 30 with B.dV or B.dV/ Na2-EDTA (7-30μg or 14-60 μg) emulsified with complete Freund?s Adjuvant and incomplete (booster). On days 14, 29, 37 blood samples were collected from the tip of the animals tail and day 45 the final bleeding and the separation of the different sera were performed. These were destined for immunoassays and neutralization assays for proteolytic, indirect hemolytic, and coagulant activity. The results of the enzyme-linked immunosorbent assay showed that both anti-B.dV and anti-B.dV/EDTA serum had high antibody titers (1/74.850 ? 1/186.150) at the end of the immunization protocol. Regarding the Western Blot, the anti-B.dV/EDTA serum recognized the main bands, corresponding to the venom proteins, in a similar way as the anti-B.dV. Additionally, the experimental sera produced showed neutralizing capacity of the main toxic activities tested in vitro. This result shows that Na2-EDTA does not affect the immunogenicity of proteins since animals immunized with B.dV/ Na2-EDTA respond to B.diporus venom in a similar way to animals immunized with venom without inhibitor.
Fil: López, Gisela Lumila. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Nordeste. Instituto de Química Básica y Aplicada del Nordeste Argentino. Universidad Nacional del Nordeste. Facultad de Ciencias Exactas Naturales y Agrimensura. Instituto de Química Básica y Aplicada del Nordeste Argentino; Argentina
Fil: Leiva, Laura Cristina Ana. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Nordeste. Instituto de Química Básica y Aplicada del Nordeste Argentino. Universidad Nacional del Nordeste. Facultad de Ciencias Exactas Naturales y Agrimensura. Instituto de Química Básica y Aplicada del Nordeste Argentino; Argentina
Fil: Fusco, Luciano Sebastian. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Nordeste. Instituto de Química Básica y Aplicada del Nordeste Argentino. Universidad Nacional del Nordeste. Facultad de Ciencias Exactas Naturales y Agrimensura. Instituto de Química Básica y Aplicada del Nordeste Argentino; Argentina
XXIII Annual Meeting of the Argentinean Biology Society
Buenos Aires
Argentina
Sociedad Argentina de Biología
Materia
Na2EDTA
ANTIVENOM
ANTIBODY
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:11336/224955

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spelling Evaluation of an experimental botropic antivenom produced with venom blocked with Na2-EDTALópez, Gisela LumilaLeiva, Laura Cristina AnaFusco, Luciano SebastianNa2EDTAANTIVENOMANTIBODYhttps://purl.org/becyt/ford/1.6https://purl.org/becyt/ford/1Snake envenoming is a serious medical problem in tropical developing countries and antivenoms are the only effective therapy. Antisera are produced by immunizing horses with snake venom and adjuvant, usually Freund´s adjuvant. Botropic venoms contain metalloproteinases (SVMPs), responsible for the local effects of envenoming, such as hemorrhage, edema and myotoxicity as well as systemic bleeding. SVMPs represent around 34,2 % of the protein composition from B. diporus venom and causing lesions during the immunization of animals. In search of new processes, focused on animal welfare for the production immunobiologicals, and taking into account that the toxic action of SVMPs is inhibited by Na2-EDTA, in the present work, the immune response of animals inoculated with B.diporus venom blocked with Na2-EDTA was evaluated. For this proposal, the B. diporus venom (1,9 mg/mL) was blocked by Na2-EDTA (200 mM, B.dV/ Na2-EDTA) and used as antigen. Previously to the inoculation, the excess of chelate was removed by molecular exclusion chromatography (Sephadex G-25). Likewise, the venom without inhibitor (B.dV) received the same treatment and in both cases, the effective neutralization of SVMPs using azocasein as substrate was determined. Group of 5 CF-1 mice were immunized subcutaneously on days 0, 15 and 30 with B.dV or B.dV/ Na2-EDTA (7-30μg or 14-60 μg) emulsified with complete Freund?s Adjuvant and incomplete (booster). On days 14, 29, 37 blood samples were collected from the tip of the animals tail and day 45 the final bleeding and the separation of the different sera were performed. These were destined for immunoassays and neutralization assays for proteolytic, indirect hemolytic, and coagulant activity. The results of the enzyme-linked immunosorbent assay showed that both anti-B.dV and anti-B.dV/EDTA serum had high antibody titers (1/74.850 ? 1/186.150) at the end of the immunization protocol. Regarding the Western Blot, the anti-B.dV/EDTA serum recognized the main bands, corresponding to the venom proteins, in a similar way as the anti-B.dV. Additionally, the experimental sera produced showed neutralizing capacity of the main toxic activities tested in vitro. This result shows that Na2-EDTA does not affect the immunogenicity of proteins since animals immunized with B.dV/ Na2-EDTA respond to B.diporus venom in a similar way to animals immunized with venom without inhibitor.Fil: López, Gisela Lumila. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Nordeste. Instituto de Química Básica y Aplicada del Nordeste Argentino. Universidad Nacional del Nordeste. Facultad de Ciencias Exactas Naturales y Agrimensura. Instituto de Química Básica y Aplicada del Nordeste Argentino; ArgentinaFil: Leiva, Laura Cristina Ana. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Nordeste. Instituto de Química Básica y Aplicada del Nordeste Argentino. Universidad Nacional del Nordeste. Facultad de Ciencias Exactas Naturales y Agrimensura. Instituto de Química Básica y Aplicada del Nordeste Argentino; ArgentinaFil: Fusco, Luciano Sebastian. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Nordeste. Instituto de Química Básica y Aplicada del Nordeste Argentino. Universidad Nacional del Nordeste. Facultad de Ciencias Exactas Naturales y Agrimensura. Instituto de Química Básica y Aplicada del Nordeste Argentino; ArgentinaXXIII Annual Meeting of the Argentinean Biology SocietyBuenos AiresArgentinaSociedad Argentina de BiologíaTech Science Press2023info:eu-repo/semantics/publishedVersioninfo:eu-repo/semantics/conferenceObjectReuniónJournalhttp://purl.org/coar/resource_type/c_5794info:ar-repo/semantics/documentoDeConferenciaapplication/pdfapplication/pdfhttp://hdl.handle.net/11336/224955Evaluation of an experimental botropic antivenom produced with venom blocked with Na2-EDTA; XXIII Annual Meeting of the Argentinean Biology Society; Buenos Aires; Argentina; 2022; 1-10327-95451667-5746CONICET DigitalCONICETenginfo:eu-repo/semantics/altIdentifier/url/https://www.biologia.org.ar/wp-content/uploads/2023/09/Biocell-47-Supl-3-2023-47_Jornada-Anual-SAB-2022.pdfinfo:eu-repo/semantics/altIdentifier/url/https://www.biologia.org.ar/jornadas/jornadas-anteriores/Internacionalinfo:eu-repo/semantics/openAccesshttps://creativecommons.org/licenses/by-nc-sa/2.5/ar/reponame:CONICET Digital (CONICET)instname:Consejo Nacional de Investigaciones Científicas y Técnicas2025-09-29T10:29:22Zoai:ri.conicet.gov.ar:11336/224955instacron: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:34982025-09-29 10:29:22.94CONICET Digital (CONICET) - Consejo Nacional de Investigaciones Científicas y Técnicasfalse
dc.title.none.fl_str_mv Evaluation of an experimental botropic antivenom produced with venom blocked with Na2-EDTA
title Evaluation of an experimental botropic antivenom produced with venom blocked with Na2-EDTA
spellingShingle Evaluation of an experimental botropic antivenom produced with venom blocked with Na2-EDTA
López, Gisela Lumila
Na2EDTA
ANTIVENOM
ANTIBODY
title_short Evaluation of an experimental botropic antivenom produced with venom blocked with Na2-EDTA
title_full Evaluation of an experimental botropic antivenom produced with venom blocked with Na2-EDTA
title_fullStr Evaluation of an experimental botropic antivenom produced with venom blocked with Na2-EDTA
title_full_unstemmed Evaluation of an experimental botropic antivenom produced with venom blocked with Na2-EDTA
title_sort Evaluation of an experimental botropic antivenom produced with venom blocked with Na2-EDTA
dc.creator.none.fl_str_mv López, Gisela Lumila
Leiva, Laura Cristina Ana
Fusco, Luciano Sebastian
author López, Gisela Lumila
author_facet López, Gisela Lumila
Leiva, Laura Cristina Ana
Fusco, Luciano Sebastian
author_role author
author2 Leiva, Laura Cristina Ana
Fusco, Luciano Sebastian
author2_role author
author
dc.subject.none.fl_str_mv Na2EDTA
ANTIVENOM
ANTIBODY
topic Na2EDTA
ANTIVENOM
ANTIBODY
purl_subject.fl_str_mv https://purl.org/becyt/ford/1.6
https://purl.org/becyt/ford/1
dc.description.none.fl_txt_mv Snake envenoming is a serious medical problem in tropical developing countries and antivenoms are the only effective therapy. Antisera are produced by immunizing horses with snake venom and adjuvant, usually Freund´s adjuvant. Botropic venoms contain metalloproteinases (SVMPs), responsible for the local effects of envenoming, such as hemorrhage, edema and myotoxicity as well as systemic bleeding. SVMPs represent around 34,2 % of the protein composition from B. diporus venom and causing lesions during the immunization of animals. In search of new processes, focused on animal welfare for the production immunobiologicals, and taking into account that the toxic action of SVMPs is inhibited by Na2-EDTA, in the present work, the immune response of animals inoculated with B.diporus venom blocked with Na2-EDTA was evaluated. For this proposal, the B. diporus venom (1,9 mg/mL) was blocked by Na2-EDTA (200 mM, B.dV/ Na2-EDTA) and used as antigen. Previously to the inoculation, the excess of chelate was removed by molecular exclusion chromatography (Sephadex G-25). Likewise, the venom without inhibitor (B.dV) received the same treatment and in both cases, the effective neutralization of SVMPs using azocasein as substrate was determined. Group of 5 CF-1 mice were immunized subcutaneously on days 0, 15 and 30 with B.dV or B.dV/ Na2-EDTA (7-30μg or 14-60 μg) emulsified with complete Freund?s Adjuvant and incomplete (booster). On days 14, 29, 37 blood samples were collected from the tip of the animals tail and day 45 the final bleeding and the separation of the different sera were performed. These were destined for immunoassays and neutralization assays for proteolytic, indirect hemolytic, and coagulant activity. The results of the enzyme-linked immunosorbent assay showed that both anti-B.dV and anti-B.dV/EDTA serum had high antibody titers (1/74.850 ? 1/186.150) at the end of the immunization protocol. Regarding the Western Blot, the anti-B.dV/EDTA serum recognized the main bands, corresponding to the venom proteins, in a similar way as the anti-B.dV. Additionally, the experimental sera produced showed neutralizing capacity of the main toxic activities tested in vitro. This result shows that Na2-EDTA does not affect the immunogenicity of proteins since animals immunized with B.dV/ Na2-EDTA respond to B.diporus venom in a similar way to animals immunized with venom without inhibitor.
Fil: López, Gisela Lumila. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Nordeste. Instituto de Química Básica y Aplicada del Nordeste Argentino. Universidad Nacional del Nordeste. Facultad de Ciencias Exactas Naturales y Agrimensura. Instituto de Química Básica y Aplicada del Nordeste Argentino; Argentina
Fil: Leiva, Laura Cristina Ana. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Nordeste. Instituto de Química Básica y Aplicada del Nordeste Argentino. Universidad Nacional del Nordeste. Facultad de Ciencias Exactas Naturales y Agrimensura. Instituto de Química Básica y Aplicada del Nordeste Argentino; Argentina
Fil: Fusco, Luciano Sebastian. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Nordeste. Instituto de Química Básica y Aplicada del Nordeste Argentino. Universidad Nacional del Nordeste. Facultad de Ciencias Exactas Naturales y Agrimensura. Instituto de Química Básica y Aplicada del Nordeste Argentino; Argentina
XXIII Annual Meeting of the Argentinean Biology Society
Buenos Aires
Argentina
Sociedad Argentina de Biología
description Snake envenoming is a serious medical problem in tropical developing countries and antivenoms are the only effective therapy. Antisera are produced by immunizing horses with snake venom and adjuvant, usually Freund´s adjuvant. Botropic venoms contain metalloproteinases (SVMPs), responsible for the local effects of envenoming, such as hemorrhage, edema and myotoxicity as well as systemic bleeding. SVMPs represent around 34,2 % of the protein composition from B. diporus venom and causing lesions during the immunization of animals. In search of new processes, focused on animal welfare for the production immunobiologicals, and taking into account that the toxic action of SVMPs is inhibited by Na2-EDTA, in the present work, the immune response of animals inoculated with B.diporus venom blocked with Na2-EDTA was evaluated. For this proposal, the B. diporus venom (1,9 mg/mL) was blocked by Na2-EDTA (200 mM, B.dV/ Na2-EDTA) and used as antigen. Previously to the inoculation, the excess of chelate was removed by molecular exclusion chromatography (Sephadex G-25). Likewise, the venom without inhibitor (B.dV) received the same treatment and in both cases, the effective neutralization of SVMPs using azocasein as substrate was determined. Group of 5 CF-1 mice were immunized subcutaneously on days 0, 15 and 30 with B.dV or B.dV/ Na2-EDTA (7-30μg or 14-60 μg) emulsified with complete Freund?s Adjuvant and incomplete (booster). On days 14, 29, 37 blood samples were collected from the tip of the animals tail and day 45 the final bleeding and the separation of the different sera were performed. These were destined for immunoassays and neutralization assays for proteolytic, indirect hemolytic, and coagulant activity. The results of the enzyme-linked immunosorbent assay showed that both anti-B.dV and anti-B.dV/EDTA serum had high antibody titers (1/74.850 ? 1/186.150) at the end of the immunization protocol. Regarding the Western Blot, the anti-B.dV/EDTA serum recognized the main bands, corresponding to the venom proteins, in a similar way as the anti-B.dV. Additionally, the experimental sera produced showed neutralizing capacity of the main toxic activities tested in vitro. This result shows that Na2-EDTA does not affect the immunogenicity of proteins since animals immunized with B.dV/ Na2-EDTA respond to B.diporus venom in a similar way to animals immunized with venom without inhibitor.
publishDate 2023
dc.date.none.fl_str_mv 2023
dc.type.none.fl_str_mv info:eu-repo/semantics/publishedVersion
info:eu-repo/semantics/conferenceObject
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http://purl.org/coar/resource_type/c_5794
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status_str publishedVersion
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dc.identifier.none.fl_str_mv http://hdl.handle.net/11336/224955
Evaluation of an experimental botropic antivenom produced with venom blocked with Na2-EDTA; XXIII Annual Meeting of the Argentinean Biology Society; Buenos Aires; Argentina; 2022; 1-1
0327-9545
1667-5746
CONICET Digital
CONICET
url http://hdl.handle.net/11336/224955
identifier_str_mv Evaluation of an experimental botropic antivenom produced with venom blocked with Na2-EDTA; XXIII Annual Meeting of the Argentinean Biology Society; Buenos Aires; Argentina; 2022; 1-1
0327-9545
1667-5746
CONICET Digital
CONICET
dc.language.none.fl_str_mv eng
language eng
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info:eu-repo/semantics/altIdentifier/url/https://www.biologia.org.ar/jornadas/jornadas-anteriores/
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