Fig. 1: Single-molecule multiplexed sensing using an electro-optical nanopore platform.

a Workflow for detection of miRNAs directly from patient serum. (i) Serum from patients was incubated with length-encoded molecular probes consisting of a DNA carrier and molecular beacon (MB). (ii) Electro-optical sensing was then performed, and (iii) the miRNA expression levels were determined. b Schematic representation of the preparation of size-coded DNA probes and their binding to respective miRNA targets. Lambda-phage DNA is enzymatically engineered to generate different size fragments with 12-base single-stranded termini. Customised MB sequences are then assigned and hybridised to these fragments. Upon binding to the corresponding miRNAs, the ‘stem-loop’ structure unfolds, resulting in an increase in fluorescence intensity induced by the increase in distance between the fluorophore and the quencher. c Representative photon and current time traces for simultaneous detection of miR-141-3p and miR-375-3p from a patient in remission and a patient with active prostate cancer. Traces were recorded using asymmetric KCl buffer conditions (inside/outside nanopipette, 40/400 mM) at −300 mV bias and a laser power of 90 ± 4 µW. Scale bar of photon trace (green): vertical, 100 counts/ms, horizontal, 2.5 s. Scale bar of current trace: vertical, 30 pA, horizontal, 2.5 s.