Laborious and costly detection of miRNAs has brought challenges to its practical applications, especially for home health care, rigorous military medicine, and the third world. In this work, we present a pH-responsive miRNA amplification method, which allows the detection of miRNA just using a pH test paper. The operation is easy and no other costly instrument is involved, making the method very friendly. In our strategy, a highly efficient isothermal amplification of miRNA is achieved using an improved netlike rolling circle amplification (NRCA) technique. Large amounts of H can be produced as a byproduct during the amplification to induce significant changes of pH, which can be monitored directly using a pH test paper or pH-sensitive indicators. The degree of color changes depends on the amount of miRNA, making it possible for quantitative analysis. As an example, the method is successfully applied to quantify a miRNA (miR-21) in cancer cells. The results agree well with that from the prevalent qRT-PCR analysis. It is the first time that a paper-based point-of-care testing (POCT) is developed for the detection of miRNAs, which might promote the popularization of miRNAs working as biomarkers for diagnostic purposes.

Abnormal expression of miRNAs always occurs in solid tumors. Thus, it is critical to sensitively and selectively detect such biomarkers for the diagnosis and prognosis of diseases. Here, we report a biosensing scheme for the determination of miRNA with triple signal amplification based on target-triggered cyclic duplex specific nuclease digestion and bridge DNA-gold nanoparticles. Electrochemical signals are recorded to present initial levels of miRNA. This method is ultrasensitive with a wide linear range of 10 to 10 M. The limit of detection is down to 6.8 aM. Moreover, the overexpression of miR-21 is confirmed in lung cancer patients by the proposed method, which is in good accordance with qRT-PCR results. In addition, the developed biosensor does not need a reverse transcription process or any thermal cycling processes. Its performance satisfies the requirement for convenient, rapid, sensitive, and specific early diagnosis of cancers. Therefore, it may have great potential utility in the near future.

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