Analysis of plasma microRNAs (miRNAs) by quantitative polymerase chain reaction (qPCR) provides a potential approach for cancer diagnosis. However, absolutely quantifying low abundant plasma miRNAs is challenging with qPCR. Digital PCR offers a unique means for assessment of nucleic acids presenting at low levels in plasma. This study aimed to evaluate the efficacy of digital PCR for quantification of plasma miRNAs and the potential utility of this technique for cancer diagnosis. We used digital PCR to quantify the copy number of plasma microRNA-21-5p (miR-21–5p) and microRNA-335–3p (miR-335–3p) in 36 lung cancer patients and 38 controls. Digital PCR showed a high degree of linearity and quantitative correlation with miRNAs in a dynamic range from 1 to 10,000 copies/μL of input, with high reproducibility. qPCR exhibited a dynamic range from 100 to 1×107 copies/μL of input. Digital PCR had a higher sensitivity to detect copy number of the miRNAs compared with qPCR. In plasma, digital PCR could detect copy number of both miR-21–5p and miR-335–3p, whereas qPCR was only able to assess miR-21–5p. Quantification of the plasma miRNAs by digital PCR provided 71.8% sensitivity and 80.6% specificity in distinguishing lung cancer patients from cancer-free subjects.

Background Molecular analysis of sputum provides a promising approach for lung cancer diagnosis, yet is limited by the difficulty in collecting the specimens from individuals who can't spontaneously expectorate sputum. Lung Flute is a small self‐powered audio device that can induce sputum by generating sound waves and vibrating in the airways of the lungs. Here we propose to evaluate the usefulness of Lung Flute for sputum sampling to assist diagnosis of lung cancer. Methods Forty‐three stage I lung cancer patients and 47 cancer‐free individuals who couldn't spontaneously cough sputum were instructed to use Lung Flute for sputum sampling. Expressions of two microRNAs, miRs‐31 and 210, were determined in the specimens by qRT‐PCR. The results were compared with sputum cytology. Results Sputum was easily collected from 39 of 43 (90.7%) lung cancer patients and 42 of 47 (89.4%) controls with volume ranges from 1 to 5 ml (median, 2.6 ml). The specimens had less than 4% oral squamous cells, indicating that sputum was obtained from low respiratory tract. Expressions of miRs‐31 and 210 in sputum were considerably higher in cancer patients than cancer‐free individuals (8.990 vs. 4.514; 0.6847 vs. 0.3317; all P <0.001). Combined use of the two miRNAs produced a significantly higher sensitivity (61.5% vs. 35.9%, P = 0.002) and a slightly lower specificity (90.5% vs. 95.2%, p = 0.03) compared with cytology for lung cancer diagnosis. Conclusion Lung Flute could potentially be useful in convenient and efficient collection of sputum for molecular diagnosis of lung cancer.

In the past few years, the application of ionic liquids (ILs) had attracted more attention of the researchers. Many studies focused on extracting active components from traditional herbals using ILs as alternative solvents so as to address the issue caused by the traditional methods for extraction of natural products (NPs) with organic chemical reagents. Through the summary of reported research work, an overview was presented for the application of ILs or IL-based materials in the extraction of NPs, including flavonoids, alkaloids, terpenoids, phenylpropanoids and so on. Here, we mainly describe the application of ILs to rich the extraction of critical bioactive constituents that were reported possessing multiple therapeutic effects or pharmacological activities, from medicinal plants. This review could shed some light on the wide use of ILs in the field of natural products chemistry to further reduce the environmental damage caused by large quantity of organic chemical reagents.

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