Here, we demonstrate a phosphorodiamidate morpholino oligos (PMO)-functionalized nanochannel biosensor for label-free detection of microRNAs (miRNAs) with ultrasensitivity and high sequence specificity. PMO, as a capture probe, was covalently anchored on the nanochannel surface. Because of the neutral character and high sequence-specific affinity of PMO, hybridization efficiency between PMO and miRNAs was enhanced, thus largely decreasing background signals and highly improving the detection specificity and sensitivity. The miRNAs detection was realized through observing the change of surface charge density when PMO/miRNAs hybridization occurred. Not only could the developed biosensor specifically discriminate complementary miRNAs (Let-7b) from noncomplementary miRNAs (miR-21) and one-base mismatched miRNAs (Let-7c), but also it could detect target miRNAs in serum samples. In addition, this nanochannel-based biosensor attained a reliable limit of detection down to 1 fM in PBS and 10 fM in serum sample, respectively. It is expected that such a new method will benefit miRNA detection in clinical diagnosis.

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Compared with free miRNAs in blood, miRNAs in exosomes have higher abundance and stability. Therefore, miRNAs in exosomes can be regarded as an ideal tumor marker for early cancer diagnosis. Here, a peptide nucleic acid (PNA)-functionalized nanochannel biosensor for the ultrasensitive and specific detection of tumor exosomal miRNAs is proposed. After PNA was covalently bound to the inner surface of the nanochannels, the detection of tumor exosomal miRNAs was achieved by the charge changes on the surface of nanochannels before and after hybridization (PNA–miRNA). Due to the neutral characteristics of PNA, the efficiency of PNA–miRNA hybridization was improved by significantly reducing the background signal. This biosensor could not only specifically distinguish target miRNA-10b from single-base mismatched miRNA but also achieve a detection limit as low as 75 aM. Moreover, the biosensor was further used to detect exosomal miRNA-10b derived from pancreatic cancer cells and normal pancreatic cells. The results indicate that this biosensor could effectively distinguish pancreatic cancer tumor-derived exosomes from the normal control group, and the detection results show good consistency with those of the quantitative reverse-transcription polymerase chain reaction method. In addition, the biosensor was used to detect exosomal miRNA-10b in clinical plasma samples, and it was found that the content of exosomal miRNA-10b in cancer patients was generally higher than that of healthy individuals, proving that the method is expected to be applied for the early diagnosis of cancer.

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The rapid extraction of high-purity nucleic acids from complex biological samples using conventional methods is complicated. Therefore, in this study, a glycine-pillar[6]arene (Gly-P6) functionalized tapered nanochannels were constructed using 32mer...