QIAsymphony PAXgene Blood ccfDNA Kit
For purification of ccfDNA from plasma using the QIAsymphony SP
For purification of ccfDNA from plasma using the QIAsymphony SP
Cat. No. / ID: 768536
The fully automated QIAsymphony PAXgene Blood ccfDNA Kit procedure generates ccfDNA yields comparable to the manual QIAamp Circulating Nucleic Acid Kit procedure (see figure High performance, comparable to QIAamp Circulating Nucleic Acid Kit). Since yield is strongly dependent of donor, individual yields may vary. Onboard measures of the QIAsymphony SP minimize cross-contamination.
The workflow begins with a centrifugation step that separates plasma from the cellular fraction of the whole blood collected into PAXgene Blood ccfDNA Tubes. The plasma is then carefully transferred into a second tube. A second centrifugation step can be carried out optionally to remove any traces of buffy coat. The pure plasma is then transferred to a third tube which is loaded into the QIAsymphony SP.
In the QIAsymphony SP, plasma proteins are digested by proteinase K while ccfDNA binds to the surface of magnetic beads (see figure The QIAsymphony SP processes samples containing magnetic particles). Depending on the chosen protocol line, predominantly small ccfDNA fragments (STA protocol line) or small and large ccfDNA fragments (LAF protocol line) are isolated. Three wash steps ensure contaminant removal. Finally, ccfDNA is eluted from the magnetic particles and is ready for use in downstream applications (see figure Two protocol lines of the QIAsymphony PAXgene Blood ccfDNA Kit for different research needs).
ccfDNA purified using the QIAsymphony PAXgene Blood ccfDNA Kit and the QIAsymphony SP is ready for use in a wide range of downstream applications, including:
Whole blood of 4 healthy donors was collected into either EDTA or PAXgene Blood ccfDNA Tubes. Blood was left at room temperature for up to 7 days. Each day, ccfDNA was isolated from samples, bisulphite converted (Epitect FAST kit, QIAGEN) and analyzed with the GEMINI assay (Clinical Genomics) based on 2 methylated (BCAT1 and IKZF1) and 1 control (ACTB) target. Three PCR replicates per target sequence were analyzed. ccfDNA from blood stored in EDTA tubes generated more false positives than from blood stored in PAXgene Blood ccfDNA Tubes (data courtesy of Clinical Genomics).
Features | Specifications |
---|---|
Applications | Wide spectrum of research applications, including analysis of cancer tumor DNA and non-invasive prenatal testing |
Format | Magnetic beads |
Input volume | 2.4 ml or 4.8 ml plasma |
Throughput | 192 samples per working day and instrument |
Time per run | 96 samples of 4.8 ml plasma in 5 h 40 min |
Processing | Manual: plasma centrifugation Automated: QIAsymphony SP |
Shelf life | 6 weeks for an open reagent cartridge |
Elution volume | 60 µl for the STA protocol line 60, 100 or 150 µl for the LAF protocol line |