Novel electrochemiluminescence resonance energy transfer biosensor driven by CRISPR-Cas12a system for ctDNA detection

Efficient energy donor-acceptor pairing and distance control are critical in resonance energy transfer-based electrochemiluminescence (ECL-RET) biosensor. In this work, a novel ECL-RET system with Ru(phen)₃²⁺ as ECL energy donor and Au nanocages as energy acceptor was constructed based on the characteristic that Ru(phen)₃²⁺ can be stably embedded into double-stranded DNA. On this basis, a homogeneous ECL biosensor based on target activated CRISPR-Cas12a system and driving ECL-RET effect was developed. The ECL biosensor has been successfully applied to the detection of circulating tumor DNA (ctDNA) and demonstrated efficient recognition of L858R mutation in ctDNA of non-small cell lung cancer patients. The presented ECL-RET biosensor achieves a wide linear detection range from 10 fM to 1 nM and an excellent detection limit as low as 3.0 fM for ctDNA due to its combination of the highly efficient and specific cleavage of the CRISPR-Cas12a system.