Electrode immobilization-free and sensitive electrochemical sensing of thrombin via magnetic nanoparticle-decorated DNA polymers

Abstract The development of sensitive and simple sensing strategies for trace proteins has received increasing attentions in the diagnosis and treatment of diseases. We describe here an electrode immobilization-free and homogeneous electrochemical method for sensitively detecting thrombin in human serums. The strategy relies on target-promoted proximity binding hybridization chain reaction (HCR) formation of magnetic nanoparticle (MNP)-decorated DNA polymers. Simultaneous binding of thrombin by two distinct aptamers increases their local concentrations and enables the proximity binding-induced strand displacement reaction, which leads to subsequent initiation of HCR between the methylene blue (MB)- and biotin-labeled hairpins into long DNA polymers. The streptavidin-modified MNPs further bind the biotin moieties to form the MNPs/DNA polymers. Subsequent accumulation of the MNPs/DNA polymers on the AuNP-deposited electrode can thus yield substantially enhanced current, due to the oxidation of the many MB labels, for highly sensitive detection of thrombin ranging from 5 pM∼50 nM with a detection limit of 1.1 pM in a simple electrode immobilization-free way. Selective interrogation of low levels of thrombin in diluted human serums was also verified, revealing its potential for convenient and ultrasensitive monitoring of a variety of protein biomarkers for disease diagnosis at the early stages.