Tetrahedral framework DNA-aptamer clicked affinity monolith with enhanced specificity for online profiling of Okadaic...

Developing high-performance aptamer-mediated affinity monolith represent a powerful strategy for highly efficient recognition and online precise target profiling (e.g. shellfish toxin). However, challenges including spontaneous aggregation and interchain entanglement of aptamers, still remain and severely impede high-specificity performance in affinity monoliths. Herein, a novel affinity interface engineering approach based on tetrahedral framework nucleic acid-aptamer conjugates (tFNA-apts) was proposed for building the brush-like tFNA-apt architecture and corresponding affinity monoliths. tFNA-apts with rigid scaffold and upright-top aptamers were clicked on polymer clusters, enabling robust structure, dispersive orientation and favourable conformational freedom of each aptamer in the brush-like architecture, therefore suppressed inter-strand entanglement and maximized target accessibility. Thanks to these merits, the prevalent defects including inter-strand entanglement or self-folding and substrate interfacial interactions of conventional single-strand aptamers (SS-apts) were addressed, thereby significantly boosting the specificity of affinity monolith for more efficient recognition. Excellent recognition performance was achieved on the tFNA-apts-clicked monolith with a low aptamer density of 152 μmol L^-1, as was just 1/5∼1/50 of that used in traditional SS-apts monoliths. By using okadaic acid (OA) as the model analyte, the recovery yield was significantly improved from 56∼71% (SS-apts monolith) to 90∼97% (tFNA-apts monolith), and even outperformed the monoliths with saturated aptamer loading in real shellfishes analysis. Applied to bivalves seafood (eg. clam, mussel and oyster), acceptable OA recovery yields in 90.01 ± 0.85% ∼ 101.36 ± 1.28% (n = 3) were achieved with a limit of detection (LOD) of 0.075 ng mL⁻¹, and the superior analytical performance to that of classical HLB cartridge was also achieved. This work posed a facile yet efficient strategy for constructing high-performance tFNA-apt affinity monoliths, enabling improved specific recognition and precise profiling of shellfish toxins.