Optimizing Dual Aptamer Binding Assays for C-Reactive Protein by Biostatistical Analysis, Three-Dimensional Molecular Simulations, and Selection on an Integrated Microfluidic Chip

The selection of aptamers with high binding affinity by using systematic evolution of ligands by exponential enrichment (SELEX) still remains a challenge. It commonly requires a tedious post-SELEX process to enhance affinity of screened aptamers. In this study, we reported a novel protocol that combined bioinformatics, molecular simulation and multiple regression analysis to identify critical parameters affecting affinity. We further utilized a new aptamer-immobilized integrated microfluidic chip to automate the characterization process and accurately validate the impact of these parameters on aptamer binding. This new approach may yield precise guidelines for aptamer selection.