Evaluation of automatic cell free DNA extraction metrics using different blood collection tubes
Abstract Liquid biopsies and cell-free DNA (cfDNA) analysis are used in numerous clinical applications. The amount of cfDNA is generally limited and many approaches require assessment of individual molecules. Optimized pre-analytical steps are therefore fundamental for accurate interpretation. Here,...
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Nature Portfolio
2025-06-01
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| Series: | Scientific Reports |
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| Online Access: | https://doi.org/10.1038/s41598-025-03508-4 |
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| author | Daniel Andersson Helena Kristiansson Manuel Luna Santamaría Huma Zafar Ivan Mijakovic Åsa Torinsson Naluai Anders Ståhlberg |
| author_facet | Daniel Andersson Helena Kristiansson Manuel Luna Santamaría Huma Zafar Ivan Mijakovic Åsa Torinsson Naluai Anders Ståhlberg |
| author_sort | Daniel Andersson |
| collection | DOAJ |
| description | Abstract Liquid biopsies and cell-free DNA (cfDNA) analysis are used in numerous clinical applications. The amount of cfDNA is generally limited and many approaches require assessment of individual molecules. Optimized pre-analytical steps are therefore fundamental for accurate interpretation. Here, we established an automated extraction approach providing cfDNA of high yield and quality. We analyzed 649 blood plasma samples collected from 23 healthy individuals and assessed the performance of four different blood collection tubes, time between sampling and plasma isolation and number of centrifugation steps. CfDNA was quantified by fluorometric analysis and quantitative polymerase chain reaction, while contaminating cellular DNA was assessed by quantitative polymerase chain reaction and parallel capillary electrophoresis. Data showed that cfDNA yield depends on both choice of blood collection tube and time between sampling and plasma isolation. Plasma isolated directly after sampling in K2EDTA tubes and plasma isolated within one week from preservative Streck tubes provided high cfDNA yield. We demonstrate that contaminating cellular DNA may be challenging to detect and that quantitative polymerase chain reaction and parallel capillary electrophoresis provide complementary information. In summary, reliable cfDNA analysis requires optimized experimental workflows, where the effects of pre-analytical factors should be considered in study designs and in clinical implementations. |
| format | Article |
| id | doaj-art-cacba9fc85a44338ae2c37b2b32cc0a1 |
| institution | DOAJ |
| issn | 2045-2322 |
| language | English |
| publishDate | 2025-06-01 |
| publisher | Nature Portfolio |
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| series | Scientific Reports |
| spelling | doaj-art-cacba9fc85a44338ae2c37b2b32cc0a12025-08-20T03:10:36ZengNature PortfolioScientific Reports2045-23222025-06-0115111310.1038/s41598-025-03508-4Evaluation of automatic cell free DNA extraction metrics using different blood collection tubesDaniel Andersson0Helena Kristiansson1Manuel Luna Santamaría2Huma Zafar3Ivan Mijakovic4Åsa Torinsson Naluai5Anders Ståhlberg6Sahlgrenska Center for Cancer Research, Department of Laboratory Medicine, Institute of Biomedicine, Sahlgrenska Academy, University of GothenburgDepartment of Clinical Genetics and Genomics, Sahlgrenska University HospitalSahlgrenska Center for Cancer Research, Department of Laboratory Medicine, Institute of Biomedicine, Sahlgrenska Academy, University of GothenburgBiobank West, Sahlgrenska University HospitalDivision of Systems and Synthetic Biology, Department of Life Sciences, Chalmers University of TechnologyBiobank West, Sahlgrenska University HospitalSahlgrenska Center for Cancer Research, Department of Laboratory Medicine, Institute of Biomedicine, Sahlgrenska Academy, University of GothenburgAbstract Liquid biopsies and cell-free DNA (cfDNA) analysis are used in numerous clinical applications. The amount of cfDNA is generally limited and many approaches require assessment of individual molecules. Optimized pre-analytical steps are therefore fundamental for accurate interpretation. Here, we established an automated extraction approach providing cfDNA of high yield and quality. We analyzed 649 blood plasma samples collected from 23 healthy individuals and assessed the performance of four different blood collection tubes, time between sampling and plasma isolation and number of centrifugation steps. CfDNA was quantified by fluorometric analysis and quantitative polymerase chain reaction, while contaminating cellular DNA was assessed by quantitative polymerase chain reaction and parallel capillary electrophoresis. Data showed that cfDNA yield depends on both choice of blood collection tube and time between sampling and plasma isolation. Plasma isolated directly after sampling in K2EDTA tubes and plasma isolated within one week from preservative Streck tubes provided high cfDNA yield. We demonstrate that contaminating cellular DNA may be challenging to detect and that quantitative polymerase chain reaction and parallel capillary electrophoresis provide complementary information. In summary, reliable cfDNA analysis requires optimized experimental workflows, where the effects of pre-analytical factors should be considered in study designs and in clinical implementations.https://doi.org/10.1038/s41598-025-03508-4Automated extractionCell-free DNAPre-analyticsLiquid biopsyBlood plasma |
| spellingShingle | Daniel Andersson Helena Kristiansson Manuel Luna Santamaría Huma Zafar Ivan Mijakovic Åsa Torinsson Naluai Anders Ståhlberg Evaluation of automatic cell free DNA extraction metrics using different blood collection tubes Scientific Reports Automated extraction Cell-free DNA Pre-analytics Liquid biopsy Blood plasma |
| title | Evaluation of automatic cell free DNA extraction metrics using different blood collection tubes |
| title_full | Evaluation of automatic cell free DNA extraction metrics using different blood collection tubes |
| title_fullStr | Evaluation of automatic cell free DNA extraction metrics using different blood collection tubes |
| title_full_unstemmed | Evaluation of automatic cell free DNA extraction metrics using different blood collection tubes |
| title_short | Evaluation of automatic cell free DNA extraction metrics using different blood collection tubes |
| title_sort | evaluation of automatic cell free dna extraction metrics using different blood collection tubes |
| topic | Automated extraction Cell-free DNA Pre-analytics Liquid biopsy Blood plasma |
| url | https://doi.org/10.1038/s41598-025-03508-4 |
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