Widespread omission of aptamer-target binding verification in aptasensor development: Consequences for sensor...

Aptamer-based biosensors (aptasensors) are widely explored for analytical and diagnostic applications because aptamers combine molecular recognition with scalable chemical synthesis. Yet aptasensors have achieved limited translational impact, motivating scrutiny of development practices. Critical reviews emphasize verifying target binding of the aptamer construct by determining its equilibrium dissociation constant (K_d, a standard measure of binding strength) prior to aptasensor development to avoid interface-contingent artifacts. We refer to this pre-development requirement as the K_d gate: an auditable, construct-matched determination of K_d in homogeneous, non-denaturing solution under sensing-relevant conditions. Here, we assess compliance with the K_d gate by analyzing 209 aptasensor publications spanning six targets. Only 20 of 209 papers reported any solution-phase verification assay, and only one passed the K_d gate; among the remaining numeric K_d reports, the reported values were weakly constrained or format-contingent and therefore not auditable. As a result, reported sensor-performance claims often cannot be interpreted, transferred, or optimized on a fully rational basis. Although practical feasibility varies across constructs and targets, the observed pattern indicates that technical difficulty alone does not explain the persistent omission. We conclude that voluntary recommendations have not, in practice, functioned as an effective gate and that passing the K_d gate should become a mandatory reporting requirement, supported by an auditable checklist for editors and reviewers.