Herein, a label-free electrochemiluminescence (ECL) aptasensor for highly sensitive determination of kanamycin was developed based on a novel "on-off-on" switch system. The first "switch on" state with remarkably high ECL intensity was obtained by the tri-layer composite films modified glassy carbon electrode towards the S2O8(2-)-O2 system. To be specific, the first layer was the Au nanoparticles decorated C60 nanoparticles (abbreviation as Au@nano-C60) as inner-layer which was prepared by the in situ generating of Au nanoparticles onto the surface of bovine serum albumin decorated nano-C60 nanoparticles. Then poly-L-histidine was first selected as a novel coreactant of S2O8(2-)-O2 system and it was adsorbed on the Au@nano-C60 modified electrode as inter-layer. Finally, a self-assembling layer of colloidal Au nanoparticles (AuNPs) was the outer-layer. The three layers were interaction by the Au-N covalent bond which could achieve a desirable initial amplified ECL signal. Successively, the capture probes (CPs) of the aptamer for the target of kanamycin was anchored on the resultant tri-layer composite films modified electrode. Later, the "switch off" state was made by the quenching effect of hemin/G-quadruplex DNAzymes towards S2O8(2-)-O2 system according to the DNA hybridization reaction of an assistant probes (APs, guanine-rich nucleic acid) with CPs which could generate a large amount of hemin/G-quadruplex DNAzymes in the presence of hemin with a simple and label-free process. As expected, the second "switch on" state was the ECL signal recovery when the target of kanamycin was present, it is attributed to that the formation of the aptamer-kanamycin complex makes the quencher of hemin/G-quadruplex DNAzymes release from the sensing interface. With the association of "on-off-on" switch system, a linear response about 9 orders of magnitude for kanamycin detection was obtained from 0.15 nM to 170 mm as well as a detection limit down to 45 pM.
Keywords: Au@nano-C(60); Electrochemiluminescence (ECL); Hemin/G-quadruplex DNAzymes; Label-free; “On-off-on” switch system.
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