Aspectos clave para el desarrollo de inmunosensores digitales electroquímicos sostenibles

Resumen

Point-of-care (POC) tests are necessary because they allow a rapid diagnosis to be made outside of a specialized laboratory. Within the POC, we can find relatively sophisticated devices capable of quantifying and storing the information provided by one or several sensors, with the disadvantage that in most cases their cost per unit is very high. Within POC, paper-based devices are extremely simple and low-cost, such as the so-called lateral flow tests, in which an immunoassay is performed with just a drop of saliva, blood, sweat, or urine. Some examples are the pregnancy test, the HIV test or test for the detection of SARS-CoV-2. The lateral-flow tests have a qualitative or semi-quantitative response that allows a visual inspection of the result, however, when a more precise and unequivocal quantification is required, it is necessary to incorporate a reading system. Different techniques have been studied to quantify the signal, the most promising being the use of electrochemical techniques. However, up to date all the proposals have been carried out from external power sources and electronic modules, breaching part of the ASSURED criteria proposed by the WHO. Therefore, there is an urgent need to find a viable alternative capable of providing a reading at a minimum cost and without requiring external equipment. Consequently, this thesis delves into various aspects aimed at studying the feasibility of a single-use self-powered electrochemical test that can meet the criteria of cost, simplicity and sensitivity required to become a global impact test. The first experimental chapter describes which effects may have the selection of different papers commonly used in paper-based analytical devices on the electrochemical response. This study is innovative as there was no detailed study on the effect of paper porosity on the electrochemical response despite the larger number of studies published to date. The results obtained experimentally were validated by means of simulations carried out in COMSOL Multyphisics®. Thanks to this, it has been possible to demonstrate that the choice of paper plays a determining role in the sensitivity of the system. The second experimental chapter, a prototype of a paper-based device is presented in which a completely self-powered immunoassay is performed for the detection of the CRP biomarker. The detection is carried out by means of antibodies marked with the HRP enzyme. Then, Mg is used as anode while benzoquinone (BQ) produced by the enzyme is used as catholyte, therefore, the energy produced is directly proportional to the concentration of the biomarker. The detection values obtained with this system are within a relevant clinical range and with output voltages greater than 1.55 V. This chapter presents a starting point towards a new generation of digital and self-powered immunoassays much simpler and more affordable than those currently present in the diagnostic market. Finally, in order to replace enzymes with inorganic compounds with greater reproducibility and robustness that allow to reduce variabilities in diagnostic devices, as well as simplify the design and lower costs, in the third experimental chapter we proceeded to study the feasibility of using ceria nanoparticles (NP) as catalytic agents. The results show us that, contrary to what has been widely reported, ceria NPs do not act as a catalyst for the oxidation reaction of a mediator, but rather behave as an oxidizing agent. This characteristic means that the number of ferrocyanide molecules oxidized per NP present in the sample is fixed and the oxidation reaction stops once all the available cerium is consumed. The characterization of these materials has shown high reproducibility even under poorly controlled storage conditions, which makes them strong candidates to replace the enzymes used in current immunoassays.