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Se implementará la metodología “flipped classroom” o clase invertida en el laboratorio de Química Analítica II del Grado en Química, con el objetivo de fomentar el aprendizaje de competencias empíricas utilizando herramientas digitales.

Rheumatoid arthritis (RA) is a systemic chronic autoimmune inflammatory disease that is characterized by the destruction of bone and production of autoantibodies such as rheumatoid factor (RF) and anticitrullinated protein antibodies (ACPAs). The high prevalence of this disease and the need of affordable tools for its early detection led us to prepare the first electrochemical immunoplatform for the simultaneous determination of four RA biomarkers, the autoantibodies: RF, anti-peptidyl-arginine deiminase enzyme (anti-PAD4), anti-cyclic citrullinated peptide (anti-CCP), and anti-citrullinated vimentin (anti-MCV). Functionalized magnetic beads (MBs) were used to immobilize the specific antigens, and sandwich-type immunoassays were implemented for the amperometric detection of the four autoantibodies, using the horseradish peroxidase (HRP)/H2O2/hydroquinone (HQ) system. The immunoplatform was applied to the determination of the biomarkers in human serum of twenty-two patients diagnosed with RA and four healthy individuals, and the results were validated against ELISA tests and the certified values., Ministerio de Ciencia e Innovacion, Comunidad de Madrid, Depto. de Química Analítica, Fac. de Ciencias Químicas, TRUE, pub

Serum level of CCL5 chemokine is considered an emerging biomarker for multiple sclerosis (MS). Due to the lack of specific assays for this disease, the development of a point-of-care test for rapid detection of MS could lead to avoiding diagnostics delays. In this paper, we report the first electrochemical immunoplatform for quantification of the CCL5 biomarker at the clinically required levels, able to discriminate between patients diagnosed with MS and healthy individuals. The immunosensing device involves protein capture from biological samples by complexation with biotinylated specific antibodies immobilized onto neutravidin-functionalized microparticles and sandwich assay with anti-CCL5 antibody and IgG labelled with horseradish peroxidase (HRP) for the enzyme-catalyzed amperometric detection of H2O2 using hydroquinone (HQ) as the redox mediator. The method shows excellent analytical performance for clinical application with a wide linear range of concentrations (0.1–300 ng·mL−1 CCL5, R2 = 0.998) and a low detection limit (40 pg·mL−1 CCL5). The biosensing platform was applied to the determination of the CCL5 endogenous content in 100-fold diluted sera both from healthy individuals and patients diagnosed with MS, with no further sample treatment in just two hours. The results were successfully compared with those obtained by the ELISA methodology., Ministerio de Ciencia, Innovacion y Universidades, Comunidad de Madrid, Depto. de Química Analítica, CAI Ciencias de la Tierra y Arqueometría, TRUE, pub

This work reports an amperometric bioplatform for the determination of anti-myelin basic protein autoantibodies (anti-MBP), a relevant biomarker for multiple sclerosis (MS) autoimmune disease. The developed configuration involves the use of carboxylated magnetic microparticles (cMBs) where the protein for specific capture of the target autoantibodies was covalently attached. The immobilized anti-MBP were further conjugated with a secondary antibody labelled with horseradish peroxidase (HRP-anti-hIgG) and amperometric transduction was performed by adding hydrogen peroxide and using hydroquinone (HQ) as redox mediator. The cathodic current resulting from the reduction of the corresponding quinone was directly proportional to the logarithmic concentration of the target autoantibodies. The analytical performance of the developed method for the determination of anti-MBP is competitive in terms of sensitivity and range of linearity with that claimed for the only biosensor reported so far in the literature, as well as with commercially available ELISA kits showing a remarkably shorter assay time. The bioplatform was applied to the analysis of serum samples of healthy individuals and patients diagnosed with MS providing results in agreement with the ELISA methodology., Ministerio de Ciencia, Innovacion y Universidades, Comunidad de Madrid, Depto. de Química Analítica, Fac. de Ciencias Químicas, TRUE, pub

This paper reports the preparation of the first dual electrochemical immunosensor for the simultaneous determination of the CXCL7 chemokine and the MMP3 metalloproteinase as relevant biomarkers for the better diagnosis and monitoring of rheumatoid arthritis derived from the multiple biomarkers measurement. The developed immunosensor involves the use of carboxylated magnetic beads (MBs) and dual screen-printed carbon electrodes (SPdCEs). Sandwich-type configurations implied the covalent immobilization of specific anti-CXCL7 (cAb1) or anti-MMP3 (cAb2) capture antibodies onto MBs and the use of biotinylated detection antibodies with further labelling with HRP-Strept conjugates. The resulting MBS bioconjugates were magnetically captured on the respective working electrode of the SPdCE and the determination of the antigens was accomplished by measuring the amperometric responses of H2O2 mediated by hydroquinone (HQ) at a potential value of −0.20 V. The dual immunosensor provided calibration plots with linear ranges between 1 and 75 ng mL−1 (CXCL7) (R2 = 0.997) and from 2.0 to 2000 pg mL−1 (MMP3) (R2 = 0.998) with detection limits of 0.8 ng mL−1 and 1.2 pg mL−1, respectively. The assay took 2 h 20 min for the simultaneous determination of both biomarkers. The dual immunosensor was successfully applied to the analysis of human serum from positive and negative RA patients., Ministerio de Ciencia, Innovación y Universidades, Comunidad de Madrid, Depto. de Química Analítica, Fac. de Ciencias Químicas, TRUE, pub

The study of the human microbiome is a multidisciplinary area ranging from the field of technology to that of personalized medicine. The possibility of using microbiota biomarkers to improve the diagnosis and monitoring of diseases (e.g., cancer), health conditions (e.g., obesity) or relevant processes (e.g., aging) has raised great expectations, also in the field of bioelectroanalytical chemistry. The well-known advantages of electrochemical biosensors—high sensitivity, fast response, and the possibility of miniaturization, together with the potential for new nanomaterials to improve their design and performance—position them as unique tools to provide a better understanding of the entities of the human microbiome and raise the prospect of huge and important developments in the coming years. This review article compiles recent applications of electrochemical (bio)sensors for monitoring microbial metabolites and disease biomarkers related to different types of human microbiome, with a special focus on the gastrointestinal microbiome. Examples of electrochemical devices applied to real samples are critically discussed, as well as challenges to be faced and where future developments are expected to go.., Ministerio de Ciencia, Innovación y Universidades (España), Comunidad de Madrid, Depto. de Química Analítica, Fac. de Ciencias Químicas, TRUE, pub

Adaptar el sistema tradicional de aprendizaje a las necesidades actuales del alumnado empleando la metodología flipped classroom en el laboratorio de Química Analítica I, con el objetivo de fomentar el aprendizaje utilizando herramientas digitales.

Chemokines are a class of cytokine whose special properties, together with their involvement and relevant role in various diseases, make them a restricted group of biomarkers suitable for diagnosis and monitoring. Despite their importance, biodetection techniques dedicated to the selective determination of one or more chemokines are very scarce. For some years now, the critical diagnosis of inflammatory diseases by detecting both cytokine and chemokine biomarkers, has had a strong impact on the development of multiple detection platforms. However, it would be desirable to implement methodologies with a higher degree of selectivity for chemokines, in order to provide more precise information. In addition, better development of biosensor technology applied to this specific field would make it possible to address the main challenges of detection methods for several diseases with a high incidence in the population, avoiding high costs and low sensitivity. Taking this into account, this review aims to present the state of the art of chemokine biodetection techniques and emphasize the role of these systems in the prevention, monitoring and treatment of various diseases associated with chemokines as a starting point for future developments that are also analyzed throughout the article., Ministerio de Ciencia, Innovación y Universidades, Comunidad de Madrid, Depto. de Química Analítica, Fac. de Ciencias Químicas, TRUE, pub

Serum level of CCL5 chemokine is considered an emerging biomarker for multiple sclerosis (MS). Due to the lack of specific assays for this disease, the development of a point-of-care test for rapid detection of MS could lead to avoiding diagnostics delays. In this paper, we report the first electrochemical immunoplatform for quantification of the CCL5 biomarker at the clinically required levels, able to discriminate between patients diagnosed with MS and healthy individuals. The immunosensing device involves protein capture from biological samples by complexation with biotinylated specific antibodies immobilized onto neutravidin-functionalized microparticles and sandwich assay with anti-CCL5 antibody and IgG labelled with horseradish peroxidase (HRP) for the enzyme-catalyzed amperometric detection of H2O2 using hydroquinone (HQ) as the redox mediator. The method shows excellent analytical performance for clinical application with a wide linear range of concentrations (0.1–300 ng·mL−1 CCL5, R2 = 0.998) and a low detection limit (40 pg·mL−1 CCL5). The biosensing platform was applied to the determination of the CCL5 endogenous content in 100-fold diluted sera both from healthy individuals and patients diagnosed with MS, with no further sample treatment in just two hours. The results were successfully compared with those obtained by the ELISA methodology., Ministerio de Ciencia, Innovación y Universidades, Comunidad de Madrid, Depto. de Química Analítica, Fac. de Ciencias Químicas, TRUE, pub

El envejecimiento es un deterioro funcional fisiológico dependiente del tiempo, que parece ser impulsado por la acumulación progresiva de defectos moleculares aleatorios en células y tejidos a lo largo de la vida. Durante el envejecimiento, se desarrolla un perfil inflamatorio crónico, estéril y de bajo grado, denominado “inflammageing”, que se confirma por el aumento de los niveles de citoquinas pro-inflamatorias y la disminución de los niveles de citoquinas anti-inflamatorias, siendo el resultado, a largo plazo, de una estimulación fisiológica crónica del sistema inmune innato, que puede ser perjudicial en la edad adulta, contribuyendo a la patogénesis de enfermedades relacionadas con la edad. El objetivo principal de esta Tesis Doctoral es el diseño, preparación y aplicación de diferentes plataformas electroquímicas inmunosensoras con características analíticas mejoradas para la determinación individual o múltiple de biomarcadores de envejecimiento en muestras clínicas. Con este propósito, se desarrollaron varias configuraciones utilizando la técnica del grafting o injertado electroquímico, ácido poli(pirrolpropiónico) y diferentes nanomateriales de carbono para la modificación de los electrodos, así como nuevas estrategias de inmovilización, y poli-HRP-Strept y nanotubos de carbono híbridos como estrategia electroactiva de amplificación de la señal. Todos los inmunosensores desarrollados se aplicaron a la determinación de diferentes biomarcadores de envejecimiento escasamente estudiados para los que no existían configuraciones similares o, cuando existían, las características analíticas de las metodologías previas se mejoraron significativamente. Todo este trabajo de investigación se llevó a cabo mediante la aplicación de la metodología habitual seguida para el diseño de inmunosensores, la elección de la estrategia de reconocimiento, la optimización de las variables experimentales, los estudios de caracterización, la construcción de la curva de calibrado y la ob, Ageing is a time-dependent physiological functional decline, which appears to be driven by the progressive accumulation through the life of a variety of random molecular defects that build up within cells and tissues. During ageing, a chronic, sterile and low-grade inflammation, called “inflammageing”, is developed as the long-term result of a chronic physiological stimulation of the innate immune system, which can become damaging in the elderly, contributing to the pathogenesis of age-related diseases. Likewise, this stage results in an inflammatory profile, which is confirmed by the high levels of pro-inflammatory cytokines and the low concentration of anti-inflammatory ones. This change mainly may be attributed to the alterations in the immune system. The main objective of this Doctoral Thesis is the design, preparation and application of different electrochemical immunosensing platforms with improved analytical characteristics for individual or multiple determination of ageing biomarkers in clinical samples. For this purpose, several configurations were developed using electrochemical grafting, poly(pyrrolepropionic) acid and different carbon nanomaterials for electrode modification, as well as novel immobilization strategies, and poly-HRP-Strept and carbon nanotubes-modified hybrids as carrier nanotags for signal amplification. All the immunosensor designed were applied to the determination of different scarcely studied “inflammageing” biomarkers for which there were no similar configurations or, when existing, the analytical characteristics of the methodology were significantly improved. All this research work was carried out by application of the usual methodology of bioelectroanalysis, which involves the design of the immunosensor and the recognition strategy, optimization of the experimental variables, characterization studies, construction of the calibration plot and establishment of the analytical characteristics, followed by the studies of repeatability

Performing bioassay formats based on enzyme and antibody recognition reactions with a single detection chip remains an unmet challenge owing to the different requirements of such bioassays. Herein, we describe a dual-marker biosensor chip, integrating enzyme and antibody-based assays for simultaneous electrochemical measurements of insulin (I) and glucose (G). Simultaneous G/I sensing has been realized by addressing key fabrication and operational challenges associated with the different assay requirements and surface chemistry. The I immunosensor relies on a peroxidase-labeled sandwich immunoassay, while G is monitored through reaction with glucose oxidase. The dual diabetes biomarker chip offers selective and reproducible detection of picomolar I and millimolar G concentrations in a single microliter sample droplet within less than 30 min, including direct measurements in whole blood and saliva samples. The resulting integrated enzymatic-immunoassay biosensor chip opens a new realm in point-of-care multiplexed biomarker detection., Leona M. and Harry B. Helmsley Charitable Trust, The Scientific and Technological Research Council of Turkey (Turkey), Ministerio de Economía y Competitividad (España), Depto. de Química Analítica, Fac. de Ciencias Químicas, TRUE, pub

Dual screen-printed carbon electrodes modified with 4-carboxyphenyl-functionalized double-walled carbon nanotubes (HOOC-Phe-DWCNTs/SPCEs) have been used as scaffolds for the preparation of electrochemical immunosensors for the simultaneous determination of the cytokines Interleukin-1b (IL-1b) and factor necrosis tumor a (TNF-a). IL-1b. Capture antibodies were immobilized onto HOOC-Phe DWCNTs/SPCEs in an oriented form making using the commercial polymeric coating Mix&Go™. Sandwich type immunoassays with amperometric signal amplification through the use of poly-HRPstreptavidin conjugates and H2O2 as HRP substrate and hydroquinone as redox mediator were implemented. Upon optimization of the experimental variables affecting the immunosensor performance, the dual immunosensor allows ranges of linearity extending between 0.5 and 100 pg/mL and from 1 to 200 pg/mL for IL-1b and TNF-a, respectively, these ranges being adequate for the determination of the cytokines in clinical samples. The achieved limits of detection were 0.38 pg/mL (IL-1b) and 0.85 pg/mL (TNF-a). In addition, the dual immunosensor exhibits excellent reproducibility of the measurements, storage stability of the anti-IL-Phe-DWCNTs/SPCE and anti-TNF-Phe-DWCNTs/SPCE conjugates, and selectivity as well as negligible cross-talking. The dual immunosensor was applied to the simultaneous determination of IL-1b and TNF-a in human serum spiked at clinically relevant concentration levels and in real saliva samples., Ministerio de Economía y Competitividad (MINECO), Comunidad de Madrid, Depto. de Química Analítica, Fac. de Ciencias Químicas, TRUE, pub