Dual-mode detection of AβO using CRISPR/Cas12a gene editing technology combined with Nanopipettes

Alzheimer's disease (AD) is a progressive and irreversible neurological disorder characterized by degeneration or dysfunction of central neurons. Recent studies reveal that soluble β-amyloid oligomers (AβO) serve as more reliable biomarkers for early AD detection. To address this issue, we specifically designed a dual-mode signal nanopipette sensor for AβO detection. The electrochemical signal is generated by ferrocene (Fc), while the fluorescence signal is based on Cy5. Upon capturing AβO, the double-stranded DNA aptamer (Apt1/DNA2) initiates a dual-signal process involving hybridization chain reaction (HCR). This mechanism promotes the formation of novel double-stranded DNA structures and activates the auxiliary cleavage function of the CRISPR/Cas12a system. In the presence of AβO, the fluorescence response exhibits linear correlation with AβO concentration (R 2  = 0.99557). The combined specificity of the double-stranded aptamer and CRISPR/Cas12a, coupled with the dual signal provided by HCR, significantly enhances the system's specificity and sensitivity. The detection method achieves a minimum detection limit of 2.15 pM with a linear detection range of 0.1–120 nM. Furthermore, this highly specific and sensitive system not only paves the way for detecting biomarkers of other diseases but also offers a highly promising research direction.