Direct electrophoretic microRNA preparation from clinical samples using nanofilter membrane
Abstract 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 nit...
| Main Authors: | , , , , , , , , |
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| Format: | Article |
| Language: | English |
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SpringerOpen
2020-01-01
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| Series: | Nano Convergence |
| Subjects: | |
| Online Access: | https://doi.org/10.1186/s40580-019-0212-3 |
| _version_ | 1818933322675388416 |
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| author | Kidan Lee Jae-Hyun Kang Hyun-Mi Kim Junhyoung Ahn Hyungjun Lim JaeJong Lee Wan-Jin Jeon Jae-Hoon Lee Ki-Bum Kim |
| author_facet | Kidan Lee Jae-Hyun Kang Hyun-Mi Kim Junhyoung Ahn Hyungjun Lim JaeJong Lee Wan-Jin Jeon Jae-Hoon Lee Ki-Bum Kim |
| author_sort | Kidan Lee |
| collection | DOAJ |
| description | Abstract 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. |
| first_indexed | 2024-12-20T04:46:33Z |
| format | Article |
| id | doaj.art-e06ea12c7eb5471a9a1a0d22d14629c0 |
| institution | Directory Open Access Journal |
| issn | 2196-5404 |
| language | English |
| last_indexed | 2024-12-20T04:46:33Z |
| publishDate | 2020-01-01 |
| publisher | SpringerOpen |
| record_format | Article |
| series | Nano Convergence |
| spelling | doaj.art-e06ea12c7eb5471a9a1a0d22d14629c02022-12-21T19:52:59ZengSpringerOpenNano Convergence2196-54042020-01-017111110.1186/s40580-019-0212-3Direct electrophoretic microRNA preparation from clinical samples using nanofilter membraneKidan Lee0Jae-Hyun Kang1Hyun-Mi Kim2Junhyoung Ahn3Hyungjun Lim4JaeJong Lee5Wan-Jin Jeon6Jae-Hoon Lee7Ki-Bum Kim8Department of Materials Science and Engineering, Seoul National UniversityDepartment of Materials Science and Engineering, Seoul National UniversityResearch Institute of Advanced Materials, Seoul National UniversityDepartment of Nano Manufacturing Technology, Nano Convergence Mechanical Systems Research Division, Korea Institute of Machinery & Materials (KIMM)Department of Nano Manufacturing Technology, Nano Convergence Mechanical Systems Research Division, Korea Institute of Machinery & Materials (KIMM)Department of Nano Manufacturing Technology, Nano Convergence Mechanical Systems Research Division, Korea Institute of Machinery & Materials (KIMM)Heimbiotek Inc.Heimbiotek Inc.Department of Materials Science and Engineering, Seoul National UniversityAbstract 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.https://doi.org/10.1186/s40580-019-0212-3Nucleic acid extractionNucleic acid preparationmiRNA extractionNanofiltrationNanoporous membraneLiquid biopsy |
| spellingShingle | Kidan Lee Jae-Hyun Kang Hyun-Mi Kim Junhyoung Ahn Hyungjun Lim JaeJong Lee Wan-Jin Jeon Jae-Hoon Lee Ki-Bum Kim Direct electrophoretic microRNA preparation from clinical samples using nanofilter membrane Nano Convergence Nucleic acid extraction Nucleic acid preparation miRNA extraction Nanofiltration Nanoporous membrane Liquid biopsy |
| title | Direct electrophoretic microRNA preparation from clinical samples using nanofilter membrane |
| title_full | Direct electrophoretic microRNA preparation from clinical samples using nanofilter membrane |
| title_fullStr | Direct electrophoretic microRNA preparation from clinical samples using nanofilter membrane |
| title_full_unstemmed | Direct electrophoretic microRNA preparation from clinical samples using nanofilter membrane |
| title_short | Direct electrophoretic microRNA preparation from clinical samples using nanofilter membrane |
| title_sort | direct electrophoretic microrna preparation from clinical samples using nanofilter membrane |
| topic | Nucleic acid extraction Nucleic acid preparation miRNA extraction Nanofiltration Nanoporous membrane Liquid biopsy |
| url | https://doi.org/10.1186/s40580-019-0212-3 |
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