Fluorescent and electrochemical bimodal bioplatform for femtomolar detection of microRNAs in blood sera

Abstract

Noncoding RNAs such as miRNAs (miRs) are extensively researched as a novel class of cancer biomarkers. Herein, we describe a new method to sensitively determine the levels miRNAs via dual signal readout involving competitive hybridization between the miR-21 target and its biotinylated analog towards the same thiolated DNA probe attached onto the surface of gold nanoparticles. Hybridization of the DNA probes by the biotinylated miRs followed by conjugation with streptavidin-horseradish peroxidase, which catalyzes the oxidation of o-phenylenediamine into 2,3-diaminophenazine, allows detecting the target using fluorescence and electrochemistry. The two signals varied in a miRNA concentration-dependent manner. The bioplatform has limits of detection of 15 fmol/L and 19 fmol/L (0.15 and 0.19 attomol in 20 μL, respectively). Furthermore, it displays a wide linear calibration range varying from 19 fmol/L to 100 pmol/L and 15 fmol/L to 100 pmol/L using voltamperometry and fluorescence, respectively. The bioplatform is able to provide very low detection limits without any DNA/RNA amplification step, an excellent selectivity toward non-complementary miRs along with an acceptable distinction with a single nucleotide mismatched sequence. Finally, the method was used to determine miR-21 expression levels in blood sera from patients diagnosed with breast cancer.