Highly repeatable detection of miRNAs in complex biological mixtures mediated by magnetically separable spherical nucleic acids and homogeneous signal amplification

Abstract

Immobilization-free photoelectrochemical (PEC) sensing holds a great potential in achieving tumor biomarker detection with high repeatability. However, most sensors suffer from weak signal and background interference, which severely limits its practical translational application toward the analysis of complex physiological samples. Herein, a homogeneous and label-free miRNA sensing system was developed by integrating magnetically separable spherical nucleic acids (SNA) and signal amplification in PEC detection. Firstly, target-capturing probes were covalently conjugated to the surface of polydopamine-coated Fe3O4 nanoparticles. SNA structures consisting of chain-extended DNA concatemers from analyte-initiated hybridization chain reactions (HCR) served as hosting reservoirs of photosensitizers (PS) to achieve signal conversion. Subsequently, magnetic separation was applied to remove complex biological matrix and PS intercalated SNA particles. The concentration signal of low abundance targets was reflected in the difference in PS concentration in the supernatant. The electron transfer on electrode surface was boosted by redox mediators in homogeneous PEC detection. Finally, the proposed nanosensing system achieved highly repeatable (relative standard deviation 2.5–3.1 %) and the anti-interference analysis of miR-21 (limit-of-detection, 0.33 fM). The practical applicability was illustrated in sensitive profiling of miR-21 in lysate of tumor cells and plasma of tumor-bearing mice.