CompELS vs SELEX: Comparative Aptamer Discovery Integrating Rational Libraries and Enzymatic Elution

Aptamers are oligonucleotides that fold into structures with target-binding activity comparable to antibodies, yet their discovery remains laborious and inconsistent. Traditional Systematic Evolution of Ligands by Exponential Enrichment (SELEX) screening often relies on randomized sequence libraries, repeated amplification, and harsh elution methods, each of which introduces bias and limits reproducibility. An alternative aptamer screening platform called Competition-Enhanced Ligand Selection (CompELS) employs repeated introduction of unenriched sequence populations while avoiding iterative elution and amplification steps. To elucidate key parameters governing selection outcomes, the effects of screening strategy, screening library design, and elution of bound DNA methodology were evaluated. A parallel study of candidate aptamer selection platforms against fluorescent protein mCherry revealed that SELEX required two additional enrichment cycles to achieve comparable enrichment of the same top 3 sequences selected via CompELS. A GT rich, empirically biased library was the predominant source of binding sequences, out-competing a tetraloop-structured library and random library. Finally, an enzymatic elution strategy enabled gentle recovery of aptamer-target complexes from either screening strategy to effectively reduce nonspecific carryover and improve the fidelity of each platform. These results demonstrate a streamlined, versatile aptamer candidate selection methodology that promises an accelerated aptamer discovery approach for protein targets and reduces bias by integrating CompELS, rational library design, and enzymatic elution.