We provided an ultrasensitive sensing strategy for microRNA detection by first employing branched DNA. With the aid of microcontact printing, we realized the multiplex sensing of different kinds of liver cancer biomarkers: microRNA and protein simultaneously. Delicately designed branched DNA included multiple complementary sticky ends as probe to microRNA capture and the double-stranded rigid branched core to increase the active sticky-ends distance and expose more DNA probes for sensitivity. The branched DNA enables 2 orders of magnitude increase in sensitivity for microRNA detection over single-stranded DNA. The limit of detection reaches as low as 10 attomolar (S/N = 3) for miR-223 and 10-12 M for α-fetoprotein. In addition, this system shows high selectivity and appropriate reproducibility (the relative standard deviation is less than 20%) in physiological media. Serum samples are tested and the results of α-fetoprotein are in good agreement with the current gold-standard method, electrochemiluminescence immunoassay analyzer. The results suggest the reliability of this approach in physiological media and show high potential in the sensing of low abundant microRNA in serum, especially for early diagnosis of primary liver cancers.
Keywords: SERS; branched DNA; frequency-shift biosensing; liver cancer biomarker; ultrasensitive detection.