A method to directly collect negatively charged nucleic acids, such as DNA and RNA, in the biosamples simply by applying an electric field in between the sample and collection buffer separated by the nanofilter membrane is proposed. The nanofilter membrane was made of low-stress silicon nitride with a thickness of 100 nm, and multiple pores were perforated in a highly arranged pattern using nanoimprint technology with a pore size of 200 nm and a pore density of 7.22 × 108/cm2. The electrophoretic transport of hsa-mir-93-5p across the membrane was confirmed in pure microRNA (miRNA) mimic solution using quantitative reverse transcription-polymerase chain reactions (qRT-PCR). Consistency of the collected miRNA quantity, stability of the system during the experiment, and yield and purity of the prepared sample were discussed in detail to validate the effectiveness of the electrical protocol. Finally, in order to check the applicability of this method to clinical samples, liquid biopsy process was demonstrated by evaluating the miRNA levels in sera of hepatocellular carcinoma patients and healthy controls. This efficient system proposed a simple, physical idea in preparation of nucleic acid from biosamples, and demonstrated its compatibility to biological downstream applications such as qRT-PCR as the conventional nucleic acid extraction protocols.

中文翻译：

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使用纳滤膜从临床样品中直接电泳制备 microRNA。


提出了一种直接收集生物样品中带负电荷的核酸（例如 DNA 和 RNA）的方法，只需在由纳滤膜分隔的样品和收集缓冲液之间施加电场即可。纳滤膜由厚度为100 nm的低应力氮化硅制成，利用纳米压印技术以高度排列的方式打出多个孔，孔径为200 nm，孔密度为7.22×108/cm2。使用定量逆转录聚合酶链反应 (qRT-PCR) 在纯 microRNA (miRNA) 模拟溶液中证实了 hsa-mir-93-5p 的跨膜电泳转运。详细讨论了收集的 miRNA 数量的一致性、实验过程中系统的稳定性以及制备样品的产量和纯度，以验证电协议的有效性。最后，为了检查该方法对临床样本的适用性，通过评估肝细胞癌患者和健康对照者血清中的miRNA水平来验证液体活检过程。这种高效的系统提出了从生物样品中制备核酸的简单物理想法，并证明了其与生物下游应用（例如作为传统核酸提取方案的 qRT-PCR）的兼容性。