High dynamic range capillary electrophoresis method for sensitive detection of low-frequency driver mutations
Abstract Cancer genomics aims to personalize treatments by identifying genetic abnormalities in cancer cells. However, current analytical techniques face limitations in simplicity and cost-effectiveness. To address these issues, we developed an enhanced capillary gel electrophoresis (CE) sequencer u...
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Nature Portfolio
2025-07-01
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| Series: | Scientific Reports |
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| Online Access: | https://doi.org/10.1038/s41598-025-01884-5 |
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| author | Nobue Tamamura Yoshihiko Hagiwara Hirokazu Kato Yusuke Ono Kenji Takahashi Kazuya Koyama Hiroki Sato Tetsuhiro Okada Hidemasa Kawabata Yu Ohtaki Chiho Maeda Miyuki Mori Shin-ichi Chiba Mishie Tanino Kenzui Taniue Takashi Anazawa Ryoji Inaba Yusuke Mizukami |
| author_facet | Nobue Tamamura Yoshihiko Hagiwara Hirokazu Kato Yusuke Ono Kenji Takahashi Kazuya Koyama Hiroki Sato Tetsuhiro Okada Hidemasa Kawabata Yu Ohtaki Chiho Maeda Miyuki Mori Shin-ichi Chiba Mishie Tanino Kenzui Taniue Takashi Anazawa Ryoji Inaba Yusuke Mizukami |
| author_sort | Nobue Tamamura |
| collection | DOAJ |
| description | Abstract Cancer genomics aims to personalize treatments by identifying genetic abnormalities in cancer cells. However, current analytical techniques face limitations in simplicity and cost-effectiveness. To address these issues, we developed an enhanced capillary gel electrophoresis (CE) sequencer using a fluorescence-acquisition technique called “HiDy” (High Dynamic range) (HiDy-CE). The HiDy-CE reduces the hardware binning region size and increases the number of regions on a charge-coupled device image sensor, expanding the dynamic range and reducing saturation risk. By applying the multi-base primer extension method to the HiDy-CE with control DNA containing known mutations, we detected variant allele frequencies (VAFs) as low as 0.5% for major KRAS hotspot mutation at codon 12 and 13. With 10 ng of DNA from small tissues obtained via fine-needle biopsy from patients with suspected pancreaticoduodenal tumors, HiDy-CE produced equivalent VAFs in KRAS compared with targeted amplicon sequencing. This demonstrated the world’s first capability of detecting mutations below 1% on CE using pathological specimens, leveraging its wide dynamic range. With only 2 ng of input DNA, the HiDy-CE provided results highly concordant with digital PCR with minimal non-specific noise. These findings underscore the HiDy-CE’s potential for sensitive detection of oncogenes such as KRAS, facilitating pre-testing before comprehensive genome profiling. |
| format | Article |
| id | doaj-art-87f584461afd42ce8226ecf23ae9e2ff |
| institution | DOAJ |
| issn | 2045-2322 |
| language | English |
| publishDate | 2025-07-01 |
| publisher | Nature Portfolio |
| record_format | Article |
| series | Scientific Reports |
| spelling | doaj-art-87f584461afd42ce8226ecf23ae9e2ff2025-08-20T03:03:40ZengNature PortfolioScientific Reports2045-23222025-07-0115111110.1038/s41598-025-01884-5High dynamic range capillary electrophoresis method for sensitive detection of low-frequency driver mutationsNobue Tamamura0Yoshihiko Hagiwara1Hirokazu Kato2Yusuke Ono3Kenji Takahashi4Kazuya Koyama5Hiroki Sato6Tetsuhiro Okada7Hidemasa Kawabata8Yu Ohtaki9Chiho Maeda10Miyuki Mori11Shin-ichi Chiba12Mishie Tanino13Kenzui Taniue14Takashi Anazawa15Ryoji Inaba16Yusuke Mizukami17Department of Advanced Genomic Community Healthcare, Asahikawa Medical UniversityNaka Diagnostic Products Division, Bio Systems Design 1st Department, Hitachi High-Tech CorporationNaka Diagnostic Products Division, Bio Systems Design 1st Department, Hitachi High-Tech CorporationDepartment of Advanced Genomic Community Healthcare, Asahikawa Medical UniversityDepartment of Advanced Genomic Community Healthcare, Asahikawa Medical UniversityDivision of Gastroenterology, Department of Medicine, Asahikawa Medical UniversityDivision of Gastroenterology, Department of Medicine, Asahikawa Medical UniversityDivision of Gastroenterology, Department of Medicine, Asahikawa Medical UniversityDivision of Gastroenterology, Department of Medicine, Asahikawa Medical UniversityDivision of Gastroenterology, Department of Medicine, Asahikawa Medical UniversityInstitute of Biomedical Research, Sapporo-Higashi Tokushukai HospitalInstitute of Biomedical Research, Sapporo-Higashi Tokushukai HospitalCenter for Advanced Research and Education, Asahikawa Medical UniversityDepartment of Diagnostic Pathology, Asahikawa Medical University HospitalDivision of Gastroenterology, Department of Medicine, Asahikawa Medical UniversityHitachi, Ltd.Naka Diagnostic Products Division, Bio Systems Design 1st Department, Hitachi High-Tech CorporationDepartment of Advanced Genomic Community Healthcare, Asahikawa Medical UniversityAbstract Cancer genomics aims to personalize treatments by identifying genetic abnormalities in cancer cells. However, current analytical techniques face limitations in simplicity and cost-effectiveness. To address these issues, we developed an enhanced capillary gel electrophoresis (CE) sequencer using a fluorescence-acquisition technique called “HiDy” (High Dynamic range) (HiDy-CE). The HiDy-CE reduces the hardware binning region size and increases the number of regions on a charge-coupled device image sensor, expanding the dynamic range and reducing saturation risk. By applying the multi-base primer extension method to the HiDy-CE with control DNA containing known mutations, we detected variant allele frequencies (VAFs) as low as 0.5% for major KRAS hotspot mutation at codon 12 and 13. With 10 ng of DNA from small tissues obtained via fine-needle biopsy from patients with suspected pancreaticoduodenal tumors, HiDy-CE produced equivalent VAFs in KRAS compared with targeted amplicon sequencing. This demonstrated the world’s first capability of detecting mutations below 1% on CE using pathological specimens, leveraging its wide dynamic range. With only 2 ng of input DNA, the HiDy-CE provided results highly concordant with digital PCR with minimal non-specific noise. These findings underscore the HiDy-CE’s potential for sensitive detection of oncogenes such as KRAS, facilitating pre-testing before comprehensive genome profiling.https://doi.org/10.1038/s41598-025-01884-5HiDyCapillary gel electrophoresisDynamic rangeLow-frequency mutationKRASFragment analysis |
| spellingShingle | Nobue Tamamura Yoshihiko Hagiwara Hirokazu Kato Yusuke Ono Kenji Takahashi Kazuya Koyama Hiroki Sato Tetsuhiro Okada Hidemasa Kawabata Yu Ohtaki Chiho Maeda Miyuki Mori Shin-ichi Chiba Mishie Tanino Kenzui Taniue Takashi Anazawa Ryoji Inaba Yusuke Mizukami High dynamic range capillary electrophoresis method for sensitive detection of low-frequency driver mutations Scientific Reports HiDy Capillary gel electrophoresis Dynamic range Low-frequency mutation KRAS Fragment analysis |
| title | High dynamic range capillary electrophoresis method for sensitive detection of low-frequency driver mutations |
| title_full | High dynamic range capillary electrophoresis method for sensitive detection of low-frequency driver mutations |
| title_fullStr | High dynamic range capillary electrophoresis method for sensitive detection of low-frequency driver mutations |
| title_full_unstemmed | High dynamic range capillary electrophoresis method for sensitive detection of low-frequency driver mutations |
| title_short | High dynamic range capillary electrophoresis method for sensitive detection of low-frequency driver mutations |
| title_sort | high dynamic range capillary electrophoresis method for sensitive detection of low frequency driver mutations |
| topic | HiDy Capillary gel electrophoresis Dynamic range Low-frequency mutation KRAS Fragment analysis |
| url | https://doi.org/10.1038/s41598-025-01884-5 |
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