Monitoring Salivary Biomarkers in Astronauts during Spaceflight: A Versatile and Reusable Portable Microfluidic Biosensor Based on Functionalized Superparamagnetic Microbeads for Performing Multiplex Chemiluminescent Immunoassays

Introduction. Portable and simple analytical devices to be used for routinary self-diagnostic applications are particularly attractive for space applications since the crewmembers must live in a microgravity environment for several months. The proposed work in the project APHRODITE (financed by Italian Space Agency) focused on the optimization of a dual-chemiluminescent immunoassay implemented onto a portable and easy-to-use platform for the quantification of two salivary hormones, cortisol and dehydroepiandrosterone (DHEA). Method. The method is based on the use of immunological techniques combined with chemiluminescent (CL) detection by exploiting a microfluidic channel integrated with a dedicated detector. Two aliquots of magnetic beads functionalized with anti-cortisol and anti-DHEA antibodies, respectively, were entrapped by magnets in two different areas along the microchannel. A solution containing the sample and the peroxidase conjugates of both DHEA and cortisol was injected to enable competition for binding the antibodies immobilized on MB surfaces. By adding the proper CL substrate, it was possible to monitor the CL signal in real time by employing an array of hydrogenated amorphous silicon (a-Si:H) photosensors. Once the assay was completed, by removing the magnets and performing the washing step, the microchannel was clean and ready to run a new assay.  Results. With this system, taking advantage of the possibility of magnetically confining the immunoprobes, chemiluminescence detectability, and photosensor sensitivity, the accurate quantification of target analytes down to 0.1 ng mL-1 for cortisol and 0.05 ng mL-1 for DHEA were obtained with high specificity and multiplexing ability.  Conclusion. Results confirmed the good detection capabilities and assay applicability of the proposed system, prompting the development of innovative universal tools used for multiplex assays that allow a health-related panel of biomarkers to be simultaneously monitored through a single analysis.