An aptamer-guided flow-cytometric biosensor for simultaneous sizing and polymer identification of nanoplastics

Nanoplastics are an emerging class of environmental contaminants whose analysis is limited by the difficulty of resolving both particle size and polymer identity at the submicrometre scale in a single high-throughput measurement. Here, a dual-parameter flow-cytometric biosensor was developed that combined polymer-selective DNA aptamers with violet side scatter (VSSC; 405 nm) to provide single-acquisition particle-size and polymer-identity readouts for nanoplastic targets. Polymer-specific aptamers against up to one-year sunlight equivalent photoaged polystyrene (PS), polyethylene terephthalate (PET), polyethylene (PE), polypropylene (PP), and polyvinyl chloride (PVC) nanoplastics were obtained by a Cell-SELEX procedure with counter-selection and were characterised by fluorescence saturation binding, giving apparent dissociation constants (K_D) of 1.19-9.27 μg mL^-1. Implemented on a CytoFLEX platform, the biosensor resolved polystyrene calibration beads down to ∼100 nm, below the practical cut-off of conventional 488 nm side scatter, and assigned polymer identity within the same VSSC window. A count-based calibration yielded limits of detection of 0.038-0.165 events μL^-1 (≈40-200 particles mL^-1), a polymer-independent figure of merit that was insensitive to fragment density or weathering state. Polymer-specific fluorescein (FITC)-channel-positive populations were retained in spiked ultrapure, tap, sea, and lake water matrices, with only modest matrix-dependent fluorescence attenuation that did not affect per-event polymer classification. The approach provided a potentially transferable biosensor architecture for nanoscale-particle analysis and was compatible with existing flow-cytometry data-analysis pipelines; adaptation to field-portable cytometers remains to be demonstrated.