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Artículo

Validation of commercial Mas receptor antibodies for utilization in Western Blotting, immunofluorescence and immunohistochemistry studies

Burghi, ValeriaIcon ; Fernandez, Natalia CristinaIcon ; Gándola, Yamila BelénIcon ; Piazza, Verónica GabrielaIcon ; Diego Tomas Quiroga; Guilhen Mario, Érica; Braga, Janaína Felix; Bader, Michael; Souza Santos, Sara Dayane; Dominici, Fernando PabloIcon ; Muñoz, Marina CeciliaIcon
Fecha de publicación: 08/2017
Editorial: Public Library of Science
Revista: Plos One
ISSN: 1932-6203
Idioma: Inglés
Tipo de recurso: Artículo publicado
Clasificación temática:
Otras Ciencias Biológicas

Resumen

Mas receptor (MasR) is a G protein-coupled receptor proposed as a candidate formediating the angiotensin (Ang)-converting enzyme 2-Ang (1-7) protective axis ofrenin-angiotensin system. Because the role of this receptor is not definitively clarified,determination of MasR tissue distribution and expression levels constitutes a criticalknowledge to fully understanding its function. Commercially available antibodies havebeen widely employed for MasR protein localization and quantification, but they havenot been adequately validated. In this study, we carried on an exhaustive evaluation offour commercial MasR antibodies, following previously established criteria. WesternBlotting (WB) and immunohistochemistry studies starting from hearts and kidneys fromwild type (WT) mice revealed that antibodies raised against different MasR domainsyielded different patterns of reactivity. Furthermore, staining patterns appearedidentical in samples from MasR knockout (MasR-KO) mice. We verified by polymerasechain reaction analysis that the MasR-KO mice used were truly deficient in thisreceptor as MAS transcripts were undetectable in either heart or kidney from thisanimal model. In addition, we evaluated the ability of the antibodies to detect thehuman c-myc-tagged MasR overexpressed in human embryonic kidney cells. Threeantibodies were capable of detecting the MasR either by WB or byimmunofluorescence, reproducing the patterns obtained with an anti c-myc antibody. Inconclusion, although three of the selected antibodies were able to detect MasR proteinat high expression levels observed in a transfected cell line, they failed to detect thisreceptor in mice tissues at physiological expression levels. As a consequence,validated antibodies that can recognize and detect the MasR at physiological levels arestill lacking.
Palabras clave: Antibodies , Mas Receptor , Immunofluorescence , Western Blotting , Angiotensin-1-7
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info:eu-repo/semantics/openAccess Excepto donde se diga explícitamente, este item se publica bajo la siguiente descripción: Creative Commons Attribution 2.5 Unported (CC BY 2.5)
Identificadores
URI: http://hdl.handle.net/11336/47229
URL: http://dx.plos.org/10.1371/journal.pone.0183278
DOI: http://dx.doi.org/10.1371/journal.pone.0183278
Colecciones
Articulos(ININFA)
Articulos de INST.DE INVEST.FARMACOLOGICAS (I)
Articulos(IQUIFIB)
Articulos de INST.DE QUIMICA Y FISICO-QUIMICA BIOLOGICAS "PROF. ALEJANDRO C. PALADINI"
Citación
Burghi, Valeria; Fernandez, Natalia Cristina; Gándola, Yamila Belén; Piazza, Verónica Gabriela; Diego Tomas Quiroga; et al.; Validation of commercial Mas receptor antibodies for utilization in Western Blotting, immunofluorescence and immunohistochemistry studies; Public Library of Science; Plos One; 12; 8; 8-2017; 1-19; e0183278
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