A rapid, sensitive, and simple-to-use biosensor for on-site detection of attomolar level microRNA biomarkers from serum extracellular vesicles

Abstract

On-site detection of microRNAs in blood-borne extracellular vesicles is important for the timely diagnosis of various diseases and physiologic conditions, including pregnancy status. This work developed a rapid, highly-sensitive capacitive biosensor to detect bovine embryonic mortality-related microRNA, miRNA-16b, in serum extracellular vesicles. The sensor was developed to achieve rapid, highly sensitive and specific detection of miRNA-16b, based on AC electrokinetically accelerated capacitive detection of DNA probe-microRNA hybridization. Using interdigitated electrodes functionalized with complementary DNA probes, this sensing method integrated capacitive measurement with simultaneous AC electrokinetic convection of analytes. Consequently, trace level microRNAs can be quantitatively detected in 30 s, with a detection limit of 22 aM from analytical buffer and 323 aM from neat serum over a linear dynamic range from 0.1 fM to 1 pM. The work also developed a simple and rapid pretreatment protocol to release miRNAs from extracellular vesicles in serum. As a result, the sensor can yield results from serum samples in minutes and diagnose embryonic mortality at day 17 of gestation with a sensitivity of 90.00% and specificity of 88.89% for clinical bovine serum samples. The sensor is fabricated from low-cost printed circuit board electrodes and is mass-production friendly. This rapid and sensitive sensor provides a foundation technology for the development of on-site diagnosis in various human and animal health applications.