Analysis of different strategies for in vitro diagnosis of human humoral biomarkers based on solid support immobilization

Abstract

The study of biomarkers and their detection in human fluids represents one of the best ways to improve the diagnosis and prevention of diseases. The use of human fluids for biomarker analysis can facilitate a friendly and non-invasive diagnosis of various pathologies that affect daily life. Early and accurate detection of these diseases is crucial, otherwise they can lead to serious conditions. In addition to its role in diagnosis, the study of biomarkers also contributes to improve treatments, as it helps to monitor response, identify new drug targets and explore therapeutic alternatives. Based on this premise, bibliographic knowledge, and previous studies of -omics technologies carried out by our research group, the present PhD thesis uses the immobilization of molecules on solid supports as a tool for the detection of biomarkers in human fluids. In this PhD thesis, we developed antibody microarrays (AbMAs) directed against human MMP-9 as a useful tool to detect this biomarker of ocular surface inflammation in tear samples. Moreover, multiplexed AbMAs were used to also detect the tear biomarkers CST4 and S100A6, providing a more complete panel of biomarkers for ocular diseases and improving patient diagnosis. In addition, we applied molecule immobilization technologies to develop a rapid test that detects collagenase activity in human synovial fluid samples by digesting an immobilized fluorescently labeled gelatin. This assay could aid in the diagnosis of diseases in which elevated extracellular matrix degradation is observed and could be used as a rapid test in primary health care. Additionally, we applied the microarray platform to study protein-protein interaction, specifically analyzing the interaction between PD-1 and PD-L1. This study demonstrates the functionality of microarrays not only in biomarker detection, but also in the analysis of protein dynamics. In conclusion, the results of the present PhD thesis deepen the existing knowledge of humoral biomarkers and provide novel approaches with potential application in the early diagnosis of diseases with an inflammatory component.