Simplified rolling circle amplification driven in situ generation of electrochemiluminescent silver nanoclusters for APE1 activity monitoring

Apurinic/apyrimidinic endonuclease 1 (APE1) plays an essential part in the base excision repair whose abnormal expression is closely correlated to the occurrence of tumors. Therefore, a sensitive assay of APE1 is crucial for the early diagnosis of cancer. Herein, we developed a simplified rolling circle amplification (RCA) strategy to drive in situ production of electrochemiluminescent (ECL) silver nanoclusters (Ag NCs) for APE1 activity monitoring. Unlike conventional RCA, which required locker DNA and padlock to form the circular template and hybridize with primer to initiate amplification, we proposed a simplified RCA that employed hairpin (HP) and padlock to directly form the circular template to initiate the amplification. Through reasonable design of the amplification templates, the RCA products were cytosine-rich (C-rich) sequences for silver ions adsorption, which were electrochemically reduced to Ag NCs. Combined with Cu-based metal-organic frameworks (Cu-MOFs) as co-reaction accelerator to facilitate the reduction of S2O82-, the ternary ECL system (Ag NCs/S2O82-/Cu-MOFs) demonstrated strong ECL intensity. The simplified RCA strategy combined with the ternary ECL sensing platform achieved the sensitive analysis of APE1 with a linear range of 1 x 10-8 to 1 x 10-2 U/mu L and a detection limit of 2.29 x 10-9 U/mu L, which presented a novel approach for sensitive monitoring of APE1 activity.