Stability of cell-free DNA from maternal plasma isolated following a single centrifugation step.
Author(s): Barrett AN, Thadani HA, Laureano-Asibal C, Ponnusamy S, Choolani M
Publication: Prenat Diagn, 2014, Vol. 34, Page 1283-8
PubMed ID: 25066782 PubMed Review Paper? No
Purpose of Paper
The purpose of this paper was to determine the effects of DNA extraction method and plasma storage duration and temperature on levels of total, long and fetal circulating cell-free DNA in plasma.
Conclusion of Paper
A higher total and fetal DNA yield was obtained using the circulating nucleic acid (CNA) kit than the QiaSymphony kit, but extraction method had no effect on the percentage of fetal DNA. There was a decrease in total and long DNA copies when plasma was stored at room temperature for 2 weeks instead of immediately recentrifuged and analyzed, but fetal DNA copy number was not affected. Storage of plasma at 4°C or -80°C did not impact total, long or fetal DNA copy number.
Studies
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Study Purpose
The purpose of this study was to determine the effects of DNA extraction method on levels of total, maternal and fetal circulating cell-free DNA in plasma. Plasma was obtained by repeat centrifugation of blood collected from 10 pregnant women carrying a male fetus in K3EDTA vacutainers. Plasma was then frozen in Lo-Bind tubes at -80°C until extraction.
Summary of Findings:
More total DNA was obtained using the CNA kit than the QiaSymphony kit (1878 versus 808.3 copies/mL plasma, p=0.0010). While more fetal DNA was obtained using the CNA kit than the QiaSymphony kit (112 versus 43.8 copies/mL, p=0.006), there was no difference in the percentage of fetal DNA between the extraction methods (10.9% by CNA and 11.9% by QiaSymphony).
Biospecimens
Preservative Types
- Frozen
Diagnoses:
- Pregnant
Platform:
Analyte Technology Platform DNA Digital PCR Pre-analytical Factors:
Classification Pre-analytical Factor Value(s) Analyte Extraction and Purification Analyte isolation method QiaSymphony extraction system
Manual CNA kit
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Study Purpose
The purpose of this study was to determine the effects of storage duration and temperature on levels of total, long and fetal circulating cell-free DNA in plasma. Blood was collected from 8 women carrying a male fetus (23-39 weeks gestation), and an additional 7 pooled blood specimens were obtained from 14 women carrying male fetuses (20-38 weeks gestation). Blood was centrifuged once and plasma was placed in fresh 1.5 mL tubes, one of which was centrifuged immediately for a second time (<4h) while the remainder were stored prior to the second centrifugation. All plasma was stored frozen in a Lo-Bind tube prior to DNA extraction.
Summary of Findings:
Storage of plasma at room temperature prior to a second centrifugation resulted in a significant increase in total circulating DNA (p=0.0068). Storage of plasma at room temperature for 2 weeks before a second centrifugation also led to an increase in the number of long copies of DNA (p=0.0145), but storage of plasma at room temperature did not affect the number of copies of fetal DNA or the percentage of fetal DNA. Storage of plasma at -80°C for up to 2 weeks or 4°C for up to 72 h prior to a second centrifugation (longest time-points examined) did not affect total, long or fetal DNA copy number.
Biospecimens
Preservative Types
- Frozen
Diagnoses:
- Other diagnoses
Platform:
Analyte Technology Platform DNA Digital PCR Pre-analytical Factors:
Classification Pre-analytical Factor Value(s) Storage Storage temperature Room temperature
4°C
-80°C
Storage Storage duration <4 h
1 day
3 days
1 week
2 weeks