Immunochemical assays belong to the most widespread methods in clinical diagnostics, combining the high specificity of antibodies and various readout techniques. Enzyme-linked immunosorbent assay (ELISA) is considered a gold standard of diagnostic assays. It is carried out in microtiter plates and utilizes enzymes (most typically horseradish peroxidase) for signal generation. Furthermore, the recent progress in nanotechnology has led to the development of various nanomaterials, which can serve as alternative immunoassay labels. For example, photon-upconversion nanoparticles (UCNPs) exhibit visible light emission upon IR excitation (anti-Stokes luminesce) and can thus be detected without optical background interference. After conjugation with biomolecules providing specific recognition, such as streptavidin or antibodies, UCNPs can serve as the detection element in upconversion-linked immunosorbent assays (ULISAs). The analytical performance of ELISA and ULISA can be enhanced by replacing the microtiter plates with magnetic micro- and nanoparticles, which combine better access to the analyte in solution with the possibility to preconcentrate the analyte in the sample using a permanent magnet.

The aim of this thesis will be the development of immunochemical assays based on magnetic particles. First, commercial magnetic microparticles will be conjugated with a model antibody (e.g., anti-human serum albumin). The conjugates will then enable the development of sandwich immunoassays based on enzyme- and UCNP-based labels. After general optimizations, the performance of magnetic immunoassays will be compared with their microtiter plate-based counterparts. Afterward, conjugates with antibodies against cancer biomarkers (e.g., prostate-specific antigen or CA-19-9) will be prepared and assay detecting the biomarker will be fully optimized. Finally, real samples of spiked serum will be analyzed, and the results will be compared to the reference approach (e.g., Roche Elecsys).