Scaffold-based molecular searching, a well-established strategy in medicinal chemistry, focuses on preserving the core structural motif of a bioactive compound while exploring variations around it to identify new molecules with retained or improved function. Drawing inspiration from this concept, I employed an analogous approach for aptamer discovery by leveraging a known functional sequence as a structural scaffold. In this design, a randomized library is strategically inserted within the predefined aptamer framework, enabling the exploration of sequence diversity while maintaining key structural features required for target binding. This strategy accelerates the identification of high-affinity aptamers for specific targets by constraining the search space to functionally relevant configurations, thereby combining the efficiency of scaffold-guided design with the versatility of combinatorial selection.

DAMPA, a metabolite of MTX generated by the rescue enzyme carboxypeptidase G2, closely resembles MTX, which leads to cross-reactivity with existing MTX aptamers and therefore limits their use in CPG2-assisted therapy monitoring. Rather than viewing this cross-reactivity as a failure of specificity, I approached it from a different perspective—from the lens of Wabi-Sabi—where beauty and utility can be found in imperfection. In this context, aptamer cross-reactivity is not inherently a problem; it is a property. The key is how we use it. By identifying a DAMPA-specific aptamer and pairing it with a cross-reactive MTX/DAMPA counterpart, I developed a differential sensing strategy that uses mathematical deconvolution to resolve the concentration of each molecule.