Paperfluidic Chip Device for Small RNA Extraction, Amplification, and Multiplexed Analysis.

Small RNAs have been considered as potential biomarkers of various human diseases. Sensitive and multiplexed determination of small RNAs with point-of-care (POC) assay would be of great significance. Herein, an integrated paperfluidic chip device for multiplexed small RNA analysis was developed for the first time. In this system, the extraction and purification of small RNA was completed through a poly(ether sulfone) (PES) paper chip without the need for centrifugation. Subsequently, a newly designed hairpin probe-exponential amplification reaction (HP-EXPAR) was directly performed within the extraction paper chip. For the simultaneous realization of multiple detection, a multilayer paper chip was designed in a foldable manner with more portability and usability. Quantum dots (QDs) were employed as signal labels, which endowed this assay with high optical detection efficiency. Moreover, magnetic sheets were introduced as an alternative method for layer stacking, not only guaranteeing adjacent layers are in contact but also facilitating the sample dispersion. With these outstanding characteristics, our platform obtained a satisfactory sensitivity range from 3 × 105 to 3 × 108 copies with a limit of 3 × 106 copies. Additionally, the multiplex small RNA analyses from various cancer cells were in good agreement with the results of the real-time polymerase chain reaction (qRT-PCR). More importantly, simultaneous analysis of two types of miRNAs from clinical tumor samples demonstrated the clinical applicability of the system. Therefore, the proposed paper-based device shows great promise for POC applications in the future.