Development of a CRISPR/Cas13a-based electrochemiluminescence biosensing strategy for sensitive detection of α-synuclein oligomers in neurodegenerative diseases

Abstract

In this study, we report a highly sensitive CRISPR/Cas13a-based electrochemiluminescence (ECL) biosensor for detecting α-synuclein oligomers, early biomarkers for neurodegenerative diseases. The system integrates aptamer recognition, T7 transcription, CRISPR/Cas13a cleavage, and EXPAR amplification. α-Synuclein binding triggers the release of the T7 promoter, leading to RNA production that activates Cas13a, initiating collateral cleavage and EXPAR, generating double-stranded DNA that interacts with [Ru(phen)₂dppz]²⁺ to produce a measurable ECL signal. The sensor achieved an ultralow detection limit of 1.025 aM with high specificity and stability. In serum samples, recovery ranged from 95.2 % to 99.8 %, demonstrating strong accuracy. No interference was observed from unrelated proteins. The biosensor showed excellent reproducibility (intra-day RSD = 0.78 %, inter-day RSD = 2.86 %) and stable performance over 14 days. Compared to other existing methods, this strategy offers superior sensitivity and comparable dynamic range, making it highly suitable for clinical use. Although the assay requires multiple steps and approximately two hours, the tradeoff is justified by its performance. This work highlights the potential of combining CRISPR/Cas13a with ECL for ultra-sensitive biomarker detection in complex samples, supporting early diagnosis and monitoring of Parkinson's and Alzheimer's disease.