Performance
PAXgene Blood ccfDNA Tubes simplify the collection and processing of whole blood for subsequent purification of circulating, cell-free DNA (ccfDNA). The tube additive is non-crosslinking, free of formaldehyde-releasing substances, stabilizes blood cells (Figure 1 and Figure 2) and prevents apoptosis (Figure 3). Consequently, using PAXgene Blood ccfDNA Tubes for whole blood collection, storage and transport helps prevent the release of intracellular DNA into plasma (Figure 4).
ccfDNA levels are stable for up to 10 days at temperatures up to 25°C, 7 days at temperatures up to 30°C, or 3 days at temperatures up to 37°C prior to processing. Do not store blood-filled tubes below 2°C.
Thus, blood samples can be transported and stored at room temperature to process plasma later (Figure 5).
Plasma generated from whole blood collected into PAXgene Blood ccfDNA Tubes and processed with the QIAsymphony PAXgene Blood ccfDNA Kit the QIAGEN QIAamp MinElute ccfDNA Kit or the QIAGEN QIAamp Circulating Nucleic Acid Kit, yields high-quality ccfDNA for downstream analytical assays, including but not limited to PCR based assays and next-generation sequencing (NGS).
Principle
The PAXgene Blood ccfDNA Tube is a plastic, closed, evacuated tube for the collection, anticoagulation, transport and storage of human whole blood samples and the stabilization of ccfDNA. The tube contains an additive that anticoagulates blood and stabilizes blood cells via a non-crosslinking stabilization solution. This helps prevent the release of intracellular DNA into the plasma and maintains constant ccfDNA levels during sample transport and storage before processing.
Procedure
The PAXgene Blood ccfDNA Tube allows collection of 10 ml whole blood into a closed, evacuated device that contains a proprietary ccfDNA stabilization additive. Using standard venipuncture technique, blood is collected into an evacuated tube that contains 1.5 mL cell stabilization additive resulting in a ratio of 0.15 ml additive per 1 mL blood. After collection, the whole blood sample can be stored for up to 10 days at temperatures up to 25°C, 7 days at temperatures up to 30°C, or 3 days at temperatures up to 37°C prior to processing. Do not store blood-filled tubes below 2°C.
After tube centrifugation, ccfDNA isolation from plasma is carried out using automated or manual ccfDNA isolation methods based on magnetic-bead or silica-membrane technologies, respectively. The remaining total cellular fraction, or buffy coat, can be used to isolate gDNA using automated or manual DNA isolation methods based on bead or silica membrane technologies, respectively (see Supplementary Protocols).
Customized protocols also allow for automated primary tube handling, eliminating the need to manually transfer plasma from PAXgene Blood ccfDNA Tubes to processing tubes (see Save Time and Resources – Primary Tube Handling Using the PAXgene Blood ccfDNA Tube).
Applications
ccfDNA purified from plasma generated from whole blood collected in PAXgene Blood ccfDNA Tubes can be used in a wide range of downstream applications, such as:
- PCR, including digital, multiplex and quantitative real-time PCR, and methylation-based assays
- Next-generation sequencing
