Multi-signal amplification electrochemiluminescence biosensor based on GO-CDTPA-AuNPs composites and PET-RAFT for ultrasensitive detection of KRAS G12C mutation

Abstract

The detection of the Kirsten Rat Sarcoma (KRAS) G12C point mutation plays an important role in the early diagnosis and individualized treatment of non-small cell lung cancer (NSCLC). Herein, a novel electrochemiluminescence (ECL) biosensor for the detection of KRAS G12C mutation via GO-CDTPA-AuNPs composites and photo-induced electron/energy transfer reversible addition-fragment chain transfer (PET-RAFT) multi-signal amplification strategy was fabricated for the first time. In this work, gold nanoparticles (AuNPs) were loaded onto the graphene oxide (GO) surface to form a luminol electrochemiluminescence enhanced composite by simple amide reaction and electrostatic adsorption. Specifically, in the presence of KRAS G12C mutation DNA, i.e. target DNA (tDNA), self-assembled hairpin DNA (hDNA) on the biosensor electrode surface was activated after hybridization with the tDNA to expose a 3′-terminal amino group, that could be further linked to GO-CDTPA-AuNPs composites. To increase luminol-based ECL, GO-CDTPA-AuNPs composites contained a lot of photo-induced electron/energy transfer reversible addition-fragment chain transfer (PET-RAFT) agent loaded onto GO surface. PET-RAFT polymerization of N-acryloxysuccinimide (NAS) monomer was initiated to form polymer chains containing multiple binding sites for luminol. Due to the amplification of luminol attachment sites, very high quantities of luminol could be introduced onto the biosensor electrode surface, resulting in significantly amplified ECL signal intensity. Under optimal biosensor fabrication conditions, a good linear relationship between ECL signal intensity and the logarithm of tDNA concentration was observed over the range of 1 fM to 10 nM with a 0.12 fM limit of detection. Moreover, this strategy exhibited high selectivity and excellent applicability for KRAS G12C mutation detection in the clinical human serum samples. Thus, the fabricated biosensor has great potential for application in the clinical diagnosis.