The Effect of In Vitro Hemolysis on Measurement of Cell-Free DNA

Nishimura, Fumitaka; Uno, Naoki; Chiang, Ping-Chia; Kaku, Norihito; Morinaga, Yoshitomo; Hasegawa, Hiroo; Yanagihara, Katsunori

doi:10.1373/jalm.2018.027953

Cited by 10 publications

(6 citation statements)

References 18 publications

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“…It is well known that WBCs start to lyse over time in the commonly used EDTA tubes, which leads to an elevated DNA background [16,20]. The color of the plasma may be used as a rough standard in some cfDNA testing laboratories to decide whether the sample should be further tested [16,21]. In our study, none of the samples from EDTA tubes, Streck tubes, and ImproGene tubes showed obvious plasma turbidity or darkening on days 0, 3, or 7.…”

Section: Improgene Tubes Control Hemolysis In Blood Samples For 7 Daysmentioning

confidence: 53%

“…One of the major technical challenges to use cfDNA as biomarkers is the ability to quantify targets at low concentrations. Any additional DNA released from nucleated cells before the separation of plasma decreases the proportion of fetal or tumor derived cfDNA in the blood [21]. Previous studies have shown that increasing the time interval between the blood draw and the separation of plasma leads to an increase in the total DNA concentration in EDTA tubes [16,18,20].…”

Section: Discussionmentioning

confidence: 99%

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Performance of ImproGene Cell-Free DNA Tubes for Stabilization and Analysis of cfDNA in Blood Samples

Luo¹,

Wang²,

Zhang³

et al. 2021

Fetal and Pediatric Pathology

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Background: With the development of liquid biopsy technology, the demand for noninvasive prenatal testing (NIPT) is increasing rapidly. The aim of the study is to evaluate the effects of different blood collection tubes on plasma cfDNA and NIPT quality control. Methods: We investigated hemolysis, cfDNA concentration, and fragment distribution within blood samples stored in EDTA, ImproGene, and Streck tubes. The effects of ImproGene and Streck tubes on NIPT quality control were evaluated. Results: The ImproGene tubes prevented the time-dependent increase of cfDNA concentration and preserved the cfDNA fragment size distribution. For NIPT quality control, there is no significant difference in cfDNA, library concentration, and fetal fraction between ImproGene and Streck tubes samples. GC content of the samples in ImproGene tubes was closer to the human genome. Conclusion: The ImproGene cfDNA tube has excellent performance and is an effective choice for storing blood samples for NIPT testing or other cfDNA analysis.

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Section: Improgene Tubes Control Hemolysis In Blood Samples For 7 Daysmentioning

confidence: 53%

Section: Discussionmentioning

confidence: 99%

Performance of ImproGene Cell-Free DNA Tubes for Stabilization and Analysis of cfDNA in Blood Samples

Luo¹,

Wang²,

Zhang³

et al. 2021

Fetal and Pediatric Pathology

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“…These findings suggest that later tubes may be preferable for subsequent liquid biopsy analysis. Although we did not specifically investigate the impact of hemolysis on follow-up ctDNA or CTC analysis, previous studies have shown that hemolysis may negatively affect liquid biopsy analysis, leading to lower ctDNA yield or issues with blood coagulation during CTC enrichment (25–28). Another interesting finding from our hemolysis analysis was that a switch from non-hemolytic to hemolytic status of the sample (11.6%), and vice versa (5.0%) is a quite frequent phenomenon.…”

Section: Discussionmentioning

confidence: 99%

Clinical application of ISO and CEN/TS standards for liquid biopsies - information everybody wants but nobody wants to pay for

Bonstingl,

Skofler,

Ulz

et al. 2023

Preprint

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BackgroundLiquid biopsies are emerging as valuable clinical biomarkers for cancer monitoring. Despite increasing clinical use, standardization remains a challenge. ISO and CEN/TS standardized workflows exist, but their integration into clinical practice is underdeveloped. We aimed to assess the applicability of ISO and CEN/TS liquid biopsy standards in a real-world clinical setting.MethodsWe evaluated 659 peripheral blood samples from advanced prostate cancer patients against ISO and CEN/TS standards and tracked all essential criteria. This included assessing tube filing level, complete timing from blood draw until storage, transport conditions, temperature control, hemolysis score and tube draw order and its effects on hemolysis.ResultsAmong 659 samples, 92.4% (609/659) met the essential criteria for ISO and CEN/TS compliance. In total 83.8% (552/659) of blood collection tubes had high fill levels above 80% of nominal filing level. In our advanced prostate cancer cohort, 12.9% (40/311) of the evaluated plasma samples were hemolytic. Within the draw order of five blood collection tubes, hemolysis did not significantly increase from tube one to five. The complete ccfDNA ISO and CTC CEN/TS workflows were completed within an average of 168 (+/- 71 min) and 248 minutes (+/- 76 min), respectively, from blood draw until storage.ConclusionsOur study demonstrates the feasibility and benefits of adhering to ISO and CEN/T standards in a clinical liquid biopsy study. ISO and CEN/TS standards revealed that hemolysis is a common phenomenon in pre-treated advanced prostate cancer patients, as we eliminated pre-analytical errors as cause.

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“…However, in addition to the destruction of red blood cells (in vitro hemolysis), white blood cells may also be physically damaged during blood collection. Distinct from erythrocytes, white blood cells contain nuclear DNA, and their lysis during in vitro hemolysis result in cellular DNA being released into the blood [ 37 ].…”

Section: Discussionmentioning

confidence: 99%

Feasibility of Cell-Free DNA Measurement from the Earlobe during Physiological Exercise Testing

et al. 2022

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Circulating, cell-free DNA (cfDNA) has been discussed as an upcoming blood-based biomarker in exercise physiology, reflecting important aspects of exercise load. cfDNA blood sampling has evolved from elaborate venous to efficient capillary sampling from the fingertips. In this study, we aimed to evaluate the principal feasibility of cfDNA blood sampling from the earlobe. Therefore, we obtained cfDNA concentrations from the fingertips, earlobe, and the antecubital vein during physiological exercise testing. Significantly higher concentrations were obtained from the earlobe compared to fingertip samples. All of the measurement methods showed good to excellent repeatability (ICCs of 0.85 to 0.93). In addition, the control experiments revealed that repeated sampling from the earlobe but not from the fingertips increased cfDNA at rest. In summary, cfDNA sampling is feasible for all sampling sources. However, at rest, cfDNA collected from the earlobe tend to increase over time in the absence of physical load, potentially limiting this sampling method.

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The Effect of In Vitro Hemolysis on Measurement of Cell-Free DNA

Cited by 10 publications

References 18 publications

Performance of ImproGene Cell-Free DNA Tubes for Stabilization and Analysis of cfDNA in Blood Samples

Performance of ImproGene Cell-Free DNA Tubes for Stabilization and Analysis of cfDNA in Blood Samples

Clinical application of ISO and CEN/TS standards for liquid biopsies - information everybody wants but nobody wants to pay for

Feasibility of Cell-Free DNA Measurement from the Earlobe during Physiological Exercise Testing

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