Highly sensitive electrochemical PDGF-BB sensor based on protein-templated split aptamer ligation reaction

Split aptamers have attracted extensive attention in sensing benefit of optimized concise structure and low-cost, but with lower binding affinity and stability. Here, we successfully engineer three pairs of split aptamers for PDGF-BB protein and develop a highly sensitive electrochemical sensor based on protein-templated split aptamer ligation reaction. The protein template directs the reassembly of split aptamers and accelerates the ligation reaction between azide and alkyne groups attached to the split fragments. The ligation yields of RSSplit1 and RSSplit2 with reserved binding sites are found to be 72.6 +/- 1.8 % and 82.5 +/- 1.3 %, respectively, while broken binding sites also obtain a ligation yield of 61.5 +/- 2.1 % for BSSplit3. We utilize the chemically modified split aptamers in place of the "capture" and "detection" antibodies used in ELISA, directly enabling the capture of electrochemically sensitive ligated aptamer to report target binding. This protein-templated ligation strategy improves the stability and sensitivity of the electrochemical sensor, with a low detection limit of 7 pM for PDGFBB. The saturated electrochemical signal is approximately 3 times that of the non-ligated system. The proposed electrochemical sensor is also successfully applied to 50 % serum due to its excellent selectivity and bioorthogonality. This strategy avoids the signal loss caused by traditional binding interruption during washing step via converting the ternary complex of aptamer/protein/aptamer to the binary complex of ligated aptamer/ protein. This research offers a general sensing platform for other protein biomarkers and, more importantly, does not rely on expensive antibodies.