Epitope-Imprinted Magnetic Nanoparticles as a General Platform for Efficient In Vitro Evolution of Protein-Binding...

Aptamers are usually created by in vitro selection using a strategy termed systematic evolution of ligands by exponential enrichment (SELEX). Although numerous SELEX alternatives with improved selection efficiency have been developed, the overall success rate of SELEX at present is still not very ideal, which remains a great obstacle to aptamer-based research and application. In this study, an efficient and facile SELEX method was developed for in vitro screening of protein-binding aptamers, applying epitope-imprinted magnetic nanoparticles (MNPs) that exhibit highly favorable binding properties as a general affinity platform. As a proof of the principle, myoglobin (Mb) and β₂-microglobulin were employed as two target proteins. Two satisfied aptamers toward each target protein, with the dissociation constant at the 10^-8 M level and cross-reactivity less than 16.5%, were selected within three rounds, taking only 1 day. A dual aptamer-based fluorescence sandwich assay was constructed using a pair of the selected aptamers. The resulting assay allowed for quantitatively detecting Mb in human serum and distinguishing acute myocardial infarction patients from healthy individuals. The epitope-imprinted MNP-based SELEX is straightforward and generally applicable for a wide range of target proteins, providing a promising aptamer selection tool for aptamer-based research and real-world applications.