"Off" to "On" Surface-Enhanced Raman Spectroscopy Platform with Padlock Probe-Based Exponential Rolling Circle Amplification for Ultrasensitive Detection of MicroRNA 155.

In this work, an "off" to "on" surface-enhanced Raman spectroscopy (SERS) platform was constructed for ultrasensitive detection of microRNA (miRNA) by using a magnetic SERS substrate (Co@C/PEI/Ag) and padlock probe-based exponential rolling circle amplification (P-ERCA) strategy. Herein, miRNA 155 could act as primers to initiate rolling circle amplification (RCA) for producing a long repeat sequence, and then the obtained DNA would be cleaved into two kinds of single-stranded DNAs in the presence of nickase. One of the DNAs can be a new primer to initiate new cycle reactions for obtaining large numbers of the other one (trigger DNA), consequently leading to an exponential amplification. On the other hand, the hairpin DNA (H1), with a Raman label (Cy5) at one end, would form a hairpin structure to make the Cy5 closer to the SERS substrates, which could produce a strong SERS signal ("on" status). Then placeholder DNA (P2) partly hybridized with H1 to open the hairpin structure making Cy5 far away from substrates with a decreased signal ("off" status). Next, the obtained trigger DNA can complement with P2 to make the Raman label reclosed to the SERS substrates with a strong SERS signal ("on" status). From this principle, the strategy could achieve the change from "off" to "on" status. This SERS strategy exhibited a wide linear range of 100 aM to 100 pM with a low detection limit of 70.2 aM, which indicated the proposed SERS platform has potential application value for ultrasensitive bioassay of miRNA.