Person:
Martín De Santos, María Del Rosario

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First Name
María Del Rosario
Last Name
Martín De Santos
URI
https://hdl.handle.net/20.500.14352/80461
Affiliation
Universidad Complutense de Madrid
Faculty / Institute
Veterinaria
Department
Nutrición y Ciencia de los Alimentos
Area
Nutrición y Bromatología
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2020 - 20247
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Author: García García, Aina × Start date: 2020 ×

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Now showing 1 - 7 of 7
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    A novel approach to produce phage single domain antibody fragments for the detection of gluten in foods
    (Food Chemistry, 2020) García García, Aina; García Lacarra, Teresa; Martín De Santos, María Del Rosario; González Alonso, María Isabel; Madrid García, Raquel
    In this study, we demonstrated the feasibility of isolating recombinant phage-antibodies against gluten from a non-immunized library of human single-domain antibodies (dAbs). Phage display technology enabled the selection of affinity probes by successive rounds of biopanning against a biotinylated synthetic peptide comprising repetitive immunogenic gluten motifs. The analysis of a wide representation of heterologous plant species corroborated that two of the isolated clones were specific to wheat, barley and rye proteins. The phage antibody selected as the most appropriate clone for the detection of gluten in foods (dAb8E-phage) was further applied in an indirect ELISA to the analysis of 50 commercial food samples. Although the limit of detection achieved did not improve those of current immunoassays, the proposed methodology could provide promising new pathways for the generation of recombinant antibodies that allow a comprehensive determination of gluten in foods, whilst replacing the need for animal immunization.
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    Development of a new recombinant antibody, selected by phage-display technology from a celiac patient library, for detection of gluten in foods
    (Current Research in Food Science, 2023) García Calvo, Eduardo Rafael; García García, Aina; Rodríguez Gómez, Santiago; Farrais, Sergio; Martín De Santos, María Del Rosario; García Lacarra, Teresa
    Gluten, a group of ethanol-soluble proteins present in the endosperm of cereals, is extensively used in the food industry due to its ability to improve dough properties. However, gluten is also associated with a range of gluten-related diseases (GRDs), such as wheat allergies, celiac disease, and gluten intolerance. The recommended treatment for GRDs patients is a gluten-free diet. To monitor adherence to this diet, it is necessary to develop gluten-detection systems in food products. Among the available methods, immunodetection systems are the most popular due to their simplicity, reproducibility, and accuracy. The aim of this study was to generate novel high-affinity antibodies against gluten to be used as the primary reactant in an enzyme-linked immunosorbent assay (ELISA) test. These antibodies were developed by constructing an immune library from mRNA obtained from two celiac patients with a high humoral response to gluten-related proteins. The resulting library (composed by 1.1x107) was subjected to selection against gliadin using phage display technology. Following several rounds of selection, the Fab-C was selected, and demonstrated good functionality in ELISA tests, presenting a limit of detection of 15 mg/kg for detection of gluten in spiked mixtures and food products. The methodology can discriminate gluten-free products according to the current legislation.
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    Project number: 262
    Diseño y desarrollo de una herramienta audiovisual para la docencia virtual de la inspección veterinaria oficial de pescados y productos de la pesca en un mercado central
    (2022) Muñoz Atienza, Estefanía; Yagüe Sánchez, Ángel; Álvarez López, Alberto; Borrero Del Pino, Juan; García García, Aina; García Lacarra, Teresa; Hernández Cruza, Pablo Elpidio; Marín Martínez, María; Martín De Santos, María Del Rosario; Dias Araújo, Carlos; Díaz Formoso, Lara; Cintas Izarra, Luis Miguel; da Silva Serra Contente de Matos, Diogo; Feito Hermida, Javier; García Calvo, Eduardo Rafael; Gómez Sala, Beatriz; Lafuente Orte, Irene; Peña Vidal, Nuria; Rodríguez Gómez, Santiago; Sevillano González, Ester; Beltrán Crespo, Antonio; Caba Manzaneque, Elia; Cabrales Miró-Granada, Ana; Cano Muñoz, Marisa; Cañizares Cooz, Daniela; Celorrio de Ochoa, David; Dorado Nuñez, Gemma; Fernández Silva, Ana; Gutiérrez de Cabiedes de Fuente, Alejandro; Márquez Bayón, Teresa; Martín Martí, Laura; Olmeda García, Patricia; Roncero Fernández, Alejandro Francisco; Sánchez Prada, Raquel; Taberneiro Auiget, Daniel
    El objetivo global de este Proyecto de Innovación Docente es la creación de vídeos explicativos como una herramienta de aprendizaje incorporada en el Campus Virtual para mejorar el estudio sobre las actividades de higiene, inspección y control alimentario que se realizan en el Mercado Central de Pescados de Mercamadrid. La creación y el empleo de estos vídeos están dirigidos, en un principio, a los estudiantes universitarios de Grado en Veterinaria que cursan la asignatura de Higiene, Inspección y Seguridad Alimentaria. En este Proyecto se han creado vídeos explicativos que tratan sobre: (i) los controles oficiales realizados por los Técnicos Superiores Veterinarios de Mercamadrid; (ii) los riesgos sanitarios asociados al consumo de pescados, crustáceos y moluscos; (iii) la frescura del pescado; (iv) el etiquetado del pescado; (v) la identificación de especies de pescado y marisco; (vi) la prevención de fraudes en la comercialización de pescados y mariscos; y (vii) la autentificación de pescados fileteados mediante técnicas de análisis.
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    Exploring Gluten Assessment in Marketed Products through a Sandwich ELISA Methodology Based on Novel Recombinant Antibodies
    (Foods, 2024) García Calvo, Eduardo Rafael; García García, Aina; Rodríguez Gómez, Santiago; Martín De Santos, María Del Rosario; García Lacarra, Teresa
    This study presents the development of a sandwich ELISA method for gluten detection in foods, using recombinant Fab antibody fragments against gliadin. The Fabs were chemically biotinylated and immobilized on streptavidin-coated plates as capture antibodies, while alkaline phosphatase-conjugated Fabs were used as detection antibodies. Four different gliadin-binding Fabs were tested and the Fab pair Fab8E-4 and Fab-C showed the best compatibility. An indirect sandwich immunoassay, using unmodified Fab8E-4 for capture and Fab-C as the detection antibody, achieved a detection limit of 26 ng/mL of gliadin, corresponding to 10 mg/kg of gluten in foods. No cross-reactivity was observed against 60 gluten-free species commonly used in the food industry. Analysis of 50 commercial products demonstrated consistent results compared to the standard method for gluten detection. The complete lack of cross-reactivity of the developed immunoassay with oat products potentially provides an advantage over other gluten detection systems.
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    Unraveling the Properties of Phage Display Fab Libraries and Their Use in the Selection of Gliadin-Specific Probes by Applying High-Throughput Nanopore Sequencing
    (Viruses, 2024) García Calvo, Eduardo Rafael; García García, Aina; Rodríguez Gómez, Santiago; Martín De Santos, María Del Rosario; García Lacarra, Teresa
    Directed evolution is a pivotal strategy for new antibody discovery, which allowed the generation of high-affinity Fabs against gliadin from two antibody libraries in our previous studies. One of the libraries was exclusively derived from celiac patients’ mRNA (immune library) while the other was obtained through a protein engineering approach (semi-immune library). Recent advances in high-throughput DNA sequencing techniques are revolutionizing research across genomics, epigenomics, and transcriptomics. In the present work, an Oxford Nanopore in-lab sequencing device was used to comprehensively characterize the composition of the constructed libraries, both at the beginning and throughout the phage-mediated selection processes against gliadin. A customized analysis pipeline was used to select high-quality reads, annotate chain distribution, perform sequence analysis, and conduct statistical comparisons between the different selection rounds. Some immunological attributes of the most representative phage variants after the selection process were also determined. Sequencing results revealed the successful transfer of the celiac immune response features to the immune library and the antibodies derived from it, suggesting the crucial role of these features in guiding the selection of high-affinity recombinant Fabs against gliadin. In summary, high-throughput DNA sequencing has improved our understanding of the selection processes aimed at generating molecular binders against gliadin.
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    Production and Characterization of Novel Fabs Generated from Different Phage Display Libraries as Probes for Immunoassays for Gluten Detection in Food
    (Foods, 2023) García Calvo, Eduardo Rafael; García García, Aina; Rodríguez, Santiago; Takkinen, Kristiina; Martín De Santos, María Del Rosario; García Lacarra, Teresa
    Gluten is the main fraction of wheat proteins. It is widely used in the food industry because of the properties that are generated in the dough, but it is also able to trigger diseases like allergies, autoimmunity processes (such as celiac disease), and intolerances in sensitized persons. The most effective therapy for these diseases is the total avoidance of gluten in the diet because it not only prevents damage but also enhances tissue healing. To ensure the absence of gluten in food products labeled as gluten-free, accurate detection systems, like immunoassays, are required. In this work, four recombinant Fab antibody fragments, selected by phage display technology, were produced and tested for specificity and accuracy against gluten in experimental flour mixtures and commercial food products. A high-affinity probe (Fab-C) was identified and characterized. An indirect ELISA test was developed based on Fab-C that complied with the legal detection limits and could be applied in the assessment of gluten-free diets.
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    Project number: 321
    Integra y aprende. Construyendo una cadena de bloques (blockchain) de la granja a la mesa
    (2023) Cambero Rodríguez, María Isabel; Aguado Ramo, Juan Antonio; Aguilar Jaime, María Victoria; Alba Rubio, Claudio; Alonso Monge, Rebeca María Del Mar; Aragón Ramirez, Alberto; Arias López, Patricia; Arias Revenga, Jorge; Bermudez González, Guillermo; Bermejo Poza, Rubén; Blanch Rojo, María; Blanco Flores, María Dolores; Blanco Montoro, Rafael José; Bonel Ayuso, Diego Paul; Borrero Del Pino, Juan; Bugeda de Bonilla, Inés; Burgía Domínguez, Angélica; Cabeza Briales, María Concepción; Cabezas Albéniz, Almudena; Calahorra Fernández, Felipe José; Castro Madrigal, Teresa; Castro Navarro, Irma; Cervantes Navarro, Isabel; Corugedo Fernández, Lucía; Cruces Díaz, Ainhoa; Díaz Formoso, Lara; Díez Romera, Mariano; Duarete Pacheco, Sofía; Fernández Álvarez, Leonides; Fernández Solís, Claudia; Fernández-Acero Bascones, Teresa; Ferreira García-Osorio, Andrea; Fraga Perucha, Nerea; Fuente Vázquez, Jesús De La; Galicia Larrea, Paula; Gamonal Martos, Miriam; García Álvarez, Andrés; García Balboa, María Del Camino; García Calvo, Eduardo Rafael; García García, Aina; García Lacarra, Teresa; García Quiroga, Sara; García Huch, Uma Jade; González González, Noelia; González Jiménez, Lucía; Haza Duaso, Ana Isabel; Herranz Domingo, Andrea; Herranz Sorribes, Carmen; Isabel Redondo, Beatriz; Jara Pérez, Josué; Jurado Escobar, Rubén; Justo Ruiz, Carolina; Lafuente Orte, Irene; López, Cindy Alejandra; López Bote, Clemente José; López Valdeolivas, Patricia; Magro Arconada, Paula; Mallavia León, Blanca; Martín Amores, Ruth; Martín De Santos, María Del Rosario; Morales Gómez, Paloma; Moreda de Figueroa, Blanca; Moreno Conde, Helena María; Muñoz Atienza, Estefanía; Olivares Moreno, Álvaro; Peña Vidal, Nuria; Pérez Cabal, María De Los Ángeles; Pérez Sen, Raquel; Prieto Suárez, María Cinta; Paniagua Roas, Alejandra; Ramis Cervantes, Ana María; Recamal Pagán, Carlota; Remiro Yagüe, Víctor; Rodríguez Fernández, Carmina; Rodríguez Gómez, Santiago; Rodríguez Peña, José Manuel; Romera Villena, Natalia; Salazar Hijosa, Raúl; Sanabria Dominguez, Nerea; Santacruz Parra, Marta; Santos Arnaiz, Carlos; Santos López, Sergio; Torrecilla Velasco, José Santiago; Velasco De Diego, Raquel; Velasco Villar, Susana; Villanueva Suárez, María José