Auto-SELEX: A Fully Automated Microfluidic Platform for Rapid Discovery of High-Affinity Aptamers

Abstract

Aptamers are synthetic single-stranded oligonucleotides that fold into defined three-dimensional structures, enabling high-affinity and highly selective target recognition, and are increasingly explored as alternatives to antibodies in diagnostics and therapeutics owing to advantages in production cost, reproducibility, thermal stability, and generation time. However, the conventional systematic evolution of ligands by exponential enrichment (SELEX)—an iterative in vitro selection process that screens large random nucleic-acid libraries for target-binding sequences—remains labor-intensive, time-consuming, and resource-demanding, often requiring specialized expertise while exhibiting poor efficiency and low success rates. Here, we present Auto-SELEX, a fully automated microfluidic platform that enables rapid discovery of ssDNA aptamers against protein targets, completing each selection round in approximately 30 min, as opposed to days or weeks typically required by conventional or competing approaches. Auto-SELEX integrates continuous-flow free-solution electrokinetic partitioning of target-bound aptamers across an artificial sieve without immobilization or iterative washing, together with plasmonic bead-based PCR using gold nanorods for rapid amplification and library regeneration with integrated purification and thermal elution. The platform enables rapid, automated, and effective aptamer discovery, substantially accelerating the SELEX workflow as demonstrated using two structurally and biochemically distinct protein targets, human immunoglobulin E (IgE, ~190 kDa) and vitronectin (~75 kDa), yielding high-affinity and target-specific DNA aptamers after only two to three selection rounds, with dissociation constants below 20  nM and 100 nM, respectively.