Encoded particle microfluidic platform for rapid multiplexed screening and characterization of aptamers against influenza A nucleoprotein.

Aptamers represent interesting bioreceptor alternatives to antibodies when developing a bioassay and are selected by the Systematic Evolution of Ligands by Exponential Enrichment (SELEX) process. After selection, an extensive characterization process is essential to verify the binding affinity and specificity of aptamer candidates, which is the most time-consuming and costly step. In this study, we assessed a new microfluidic platform, namely Evalution™, as a rapid and high throughput aptamer characterization platform. To do this, we first selected aptamers against influenza A nucleoprotein (infA NP) by performing magnetic bead-based SELEX. The selected aptamer candidates were subsequently screened using Evalution™ for their binding kinetics and specificity towards infA NP. All aptamers showed dissociation constants (K) in the low nanomolar range (from 13 to 41 nM), and differential binding behavior towards control proteins, such as BSA and influenza B nucleoprotein (infB NP). Among 5 selected candidates, one aptamer (NP5) exhibited a significant discrimination between infA NP and infB NP and was further used to benchmark the kinetic analysis of Evalution™ (K = 41 nM) with an SPR platform (K = 17 nM). These results suggested that NP5 has the potential to be used for developing sensitive and infA NP specific aptamer-based assay. Moreover, the presented platform proved to be an efficient aptamer characterization tool for performing typical aptamer characterization experiments like binding kinetics (due to the real-time monitoring feature) and specificity assessment in a high-throughput manner due to the multiplexing capacity.