DNA nanocage-assisted multiplex fluorescence quantification of size-fractionated low-mass soluble amyloid-β oligomers

Low-mass soluble amyloid-β oligomers (LSAβOs) are increasingly recognized as key neurotoxic species in Alzheimer's disease, yet their structural heterogeneity and low abundance hinder fraction-specific quantitative analysis. Herein, we report a DNA nanocage-assisted multiplex fluorescence analytical platform for size-fractionated detection of LSAβOs in cerebrospinal fluid (CSF). The platform integrates size-selective nanocage recognition with a coupled catalytic hairpin assembly (CHA)-DNAzyme signal amplification architecture. Three customizable DNA nanocages with distinct cavity dimensions act as molecular size sieves, enabling differential capture of LSAβO fractions enriched in <10 kD, 10-30 kD, and 30-50 kD species. The released trigger strands initiate CHA-DNAzyme amplification, generating fluorescence signals that are subsequently resolved via signal deconvolution to yield apparent fraction-specific concentrations. The method achieves detection limits of 5 and 10 pM (S/N = 3) and demonstrates satisfactory recovery in spiked diluted CSF samples. This work establishes an analytical strategy that advances DNA nanocage-based sensing from size-selective recognition toward multiplex amplified quantitative profiling of heterogeneous amyloid oligomers.