Roya Tavallaie 1, 2, 3, D. Brynn Hibbert 1, Joshua McCarroll 4, Maria Kavallaris 3, 4, J. Justin Gooding 1,2,3*

1School of Chemistry, 
2Australian Centre for NanoMedicine,
3ARC Centre of Excellence in Convergent Bio-Nano Science and Technology
4Children’s Cancer Institute
The University of New South Wales
Sydney, NSW, Australia

   The ability to detect ultralow low levels of circulating microRNAs1 directly in bodily fluids is important for developing liquid biopsies, potentially replacing tumor-tissue biopsies in certain diagnostic applications. Although several methods2-4 has been developed to detect microRNA, none of them offer detecting ultralow levels of biomarker directly in patient blood. The direct analysis of patient samples eliminates the bias in the pool of sequences that are isolated, minimizes sample loss and provides a more rapid and versatile assay. Using DNA as a recognition element, we show that electrically assisted dynamic assembly of a heterogeneous network of hybridized and unhybridized probe DNA modified gold-coated magnetic nanoparticles can be used to create ultrasensitive microRNA biosensors. The sensor is the first to enable detecting as low as 10 aM microRNA concentrations in 50% blood. It produces results that are consistent with PCR, but it is functional when unpurified serum and 50% blood is used as the sample type. The analysis time required, which can be as short as 30 minutes, is clearly more rapid than several hours required for PCR. The sensor produces results within a wide dynamic range, covering 8 orders of analyte concentration from 1 nM to 10 aM. Finally, the electrical readout used in this sensor will allow a straightforward automation of the approach and a lower cost per test. 

1 Mitchell, P.S. et al. Proceedings of the National Academy of Sciences of the United States of Americ 2008, 105, 10513-10518. Circulating microRNAs as stable blood-based markers for cancer detection.
2 Miao, P., Wang, B.D., Meng, F.Y., Yin, J. & Tang, Y.G. Bioconjugate Chemistry 2015, 26, 602-607. Ultrasensitive Detection of MicroRNA through Rolling Circle Amplification on a DNA Tetrahedron Decorated Electrode. 
3 Labib, M. et al. Journal of the American Chemical Society 2013, 135, 3027-3038. Three-Mode Electrochemical Sensing of Ultralow MicroRNA Levels. 
4 Cardoso, A.R., Moreira, F.T.C., Fernandes, R. & Sales, M.G.F. Biosensors & Bioelectronics 2016, 80, 621-630. Novel and simple electrochemical biosensor monitoring attomolar levels of miRNA-155 in breast cancer.