PAXgene Blood DNA Kit
For purification of up to 500 µg genomic DNA from whole blood
- Blood collection and purification in one system
- Easy handling
- Enhanced workflow efficiency
The PAXgene Blood DNA system consists of PAXgene Blood DNA Tubes for blood collection and stabilization, and the PAXgene Blood DNA Kit for DNA purification in a single-tube procedure.
The PAXgene Blood DNA System is an integrated and standardized system for collection and stabilization of whole blood specimens and isolation of their genomic DNA. The system requires the combined use of PAXgene Blood DNA Tubes for blood collection and stabilization, followed by DNA isolation using the PAXgene Blood DNA Kit.
Blood is collected into PAXgene Blood DNA Tubes, which contain a proprietary blend of reagents that both prevents blood coagulation and stabilizes white blood cells. Blood samples can be stored in the tubes for up to 14 days at room temperature. DNA is isolated from the tubes using appropriate buffers supplied in the PAXgene Blood DNA Kit. Highly pure genomic DNA is obtained, suitable for use in a wide range of downstream applications.
The PAXgene Blood DNA procedure is shown in the flowchart " PAXgene Blood DNA procedure". Blood samples (8.5 ml) are collected in PAXgene Blood DNA Tubes, where they may be stored or transported. For DNA isolation, the blood is transferred to processing tubes (supplied already filled with cell lysis buffer), and the solution is mixed to lyse red and white blood cells. Cell nuclei and mitochondria are pelleted by centrifugation, washed, and resuspended in digestion buffer. Protein contaminants are removed by incubation with a protease. DNA is precipitated in isopropanol, washed in 70% ethanol, dried, and resuspended in resuspension buffer.
The DNA can be used in a wide range of downstream applications, including:
|applications||PCR, qPCR, real-time PCR, blotting, genotyping|
|timeperrun||60 min/8 samples|
|format||Processing tube (salting out method)|
Buffer BG4 consists of 10 mM Tris·Cl, pH 8.5.
No. DNA purified with the PAXgene Blood DNA System is free of detectable RNA contamination. In contrast to commercial salting out methods, additional RNase digestion steps are not necessary.
See subsequent FAQs for information about storage interval and temperature. Each laboratory should validate shipping procedures.
Samples in PAXgene Blood DNA Tubes can be stored at room temperature (18–25°C) for up to 14 days after blood collection.
It is strongly advised to follow the recommendations for preparing sample material provided in the corresponding Protocol Sheet to ensure reliable results.
Plasma: It is recommended to perform plasma separation immediately after blood collection when using EDTA or citrate as anticoagulant to prevent the release of genomic DNA into the plasma fraction.
Urine: Because circulating cell-free DNA in non-stabilized urine samples is rapidly degraded after sample collection due to high nuclease activity, eluates may contain no DNA or exhibit low DNA concentration. Therefore, it is recommended to stabilize urine samples. Even when using stabilized urine, it is recommended to perform a centrifugation step immediately after stabilization to prevent the release of genomic DNA from cells. Alternatively, non-stabilized urine samples can be processed immediately after collection and centrifugation using ATL-pretreatment and automated DNA extraction as described in the corresponding Protocol Sheet.
The typical A260/A280 ratio is from 1.7 to 1.9. The DNA molecules are up to 200 kb in size, with fragments of 50–150 kb predominating.
Yes, samples in PAXgene Blood DNA Tubes can be stored at 30°C for up to 3 days after blood collection. However, it is not recommended to store blood samples at temperatures higher than 30°C and storage at high temperatures should not exceed 3 days.
The typical A260/A280 ratio is from 1.7 to 1.9.
Yield of genomic DNA varies depending on the white blood cell count of the patient blood sample, the extraction method, and the elution volume. However, DNA yield typically ranges between 16–135 µg/ml whole blood. Specifically, DNA yields from magnetic bead-based extraction on the QIAsymphony ranges between 16.9–51.7 µg/ml whole blood, from silica membrane-based extraction on the QIAcube 17.1–45.8 µg/ml whole blood, and from salting-out precipitation 4.9–53.7 µg/ml whole blood.
Yes, mitochondrial and genomic DNA are both isolated.
No. The volume of Buffer BG1 (lysis buffer) prefilled in the processing tubes, and all the other buffer volumes are optimized for use with blood from one PAXgene Blood DNA Tube.
To date, DNA purified from blood samples in PAXgene Blood DNA Tubes has been successfully tested for use in PCR, including multiplex PCR.
DNA purified using the PAXgene Blood DNA System should be dissolved in Buffer BG4 and stored at 2–8°C. For long-term storage, DNA can be stored at –20°C but multiple freezing and thawing cycles should be avoided.
Yes. After dissolving the nuclei pellet in digestion buffer, Buffer BG3 (step 7), samples can be stored up to 7 days at 2–8°C or for one day at room temperature (18–25°C). The protocol can also be interrupted after adding isopropanol (step 10).
So far, we have shown that DNA purified using the PAXgene Blood DNA System can be used in PCR, including long range and multiplex PCR, restriction digestion and Southern analysis, as well as in various SNP detection procedures including,TaqMan®, LightCycler and Masscode technologies.
Do not add bleach or acidic solutions directly to the sample preparation waste. CAUTION: The sample preparation waste contains guanidinium hydrochloride from Buffer BG3, which can form highly reactive compounds when combined with bleach. For more information please consult the appropriate material safety data sheets (MSDS).
The system is designed for use only with human blood samples.
Always dispose of potentially biohazardous solutions according to your institution’s waste-disposal guidelines. Although the lysis and binding buffers in QIAamp, DNeasy, and RNeasy kits contain chaotropic agents that can inactivate some biohazardous material, local regulations dictate the proper way to dispose of biohazards. DO NOT add bleach or acidic solutions directly to the sample-preparation waste. Guanidine hydrochloride in the sample-preparation waste can form highly reactive compounds when combined with bleach.
Please access our Material Safety Data Sheets (MSDS) online for detailed information on the reagents for each respective kit.