Broad-spectrum aptamers: recent advances in selection strategies, structural mechanisms, and biosensing applications.

Broad-spectrum aptamers are single-stranded DNA or RNA molecules capable of recognizing multiple structurally or functionally related targets through adaptable molecular interactions. The increasing demand for rapid, efficient, and multi-analyte detection in complex analytical systems has exposed the limitations of conventional single-target aptamers, particularly their dependence on multiple individual recognition elements and their limited adaptability toward related analytes. This review systematically summarizes recent advances in broad-spectrum aptamers from the perspectives of target selection, structural engineering, and biosensing applications, with particular emphasis on the transition from empirical multi-target selection toward rational and mechanism-guided design. We first examine major selection strategies, including mixed-target, parallel and sequential, alternating-target, combined selection, and emerging interdisciplinary approaches, highlighting how selection pressure and target configuration influence recognition breadth, affinity, and selectivity. We then discuss structural engineering strategies, including secondary-structure regulation, chemical modification, sequence truncation and mutation, molecular simulation, and structural analysis-guided optimization, with emphasis on the molecular mechanisms underlying conformational adaptability and cross-target recognition. Recent advances in electrochemical, fluorescence, colorimetric, surface-enhanced Raman scattering, chemiluminescent, and multimodal aptasensors are further summarized, together with emerging approaches such as sensor arrays and multi-signal calibration for improving quantitative reliability in multi-analyte analysis. Finally, the major challenges and future opportunities are discussed, particularly the need to balance recognition breadth, affinity uniformity, structural adaptability, and quantitative accuracy. The integration of rational SELEX design, structural characterization, computational intelligence, nanotechnology, and miniaturized sensing platforms is expected to facilitate the development of robust, portable, and quantitatively reliable broad-spectrum aptasensors for applications in food safety, environmental monitoring, pharmaceutical analysis, and clinical diagnostics.