QIAsymphony PAXgene Blood ccfDNA Kit

For purification of ccfDNA from plasma using the QIAsymphony SP

Optimized chemistry and input volumes for PAXgene Blood ccfDNA Tubes
Separate protocol lines for small (STA) and large (LAF) ccfDNA fragments
Prefilled reagent cartrdiges with bar code for safety and ease of use
Standardized processing for high reproducibility

The PAXgene Blood ccfDNA System consists of a blood collection tube (PAXgene Blood ccfDNA Tube) and a ccfDNA purification kit for the QIAsymphony (QIAsymphony PAXgene Blood ccfDNA Kit).

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Cat No./ID: 768536

QIAsymphony PAXgene Blood ccfDNA Kit (192)

$4,060.00

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Reagent cartridges, accessories and proteinase K vials for 192 preps

For Research Use Only. Not for use in diagnostics procedures. No claim or representation is intended to provide information for the diagnosis, prevention, or treatment of a disease.

Product Details

High performance, comparable to QIAamp Circulating Nucleic Acid Kit.

Results from two studies assessing the performance of the QIAsymphony PAXgene Blood ccfDNA Kit. Study A compares ccfDNA yield and portion of double-stranded DNA of eluates generated with the QIAsymphony PAXgene Blood ccfDNA Kit to eluates from the manual QIAamp Circulating Nucleic Acid Kit. Study B summarizes performance parameters for the PAXgene ccfDNA system. The QIAsymphony PAXgene Blood ccfDNA Kit provides consistently accurate results.

PCR results from blood collected in PAXgene Blood ccfDNA Tubes remain stable over storage time.

The change in delta-C_T values of target versus control signal for ccfDNA isolated from whole blood stored in EDTA and PAXgene Blood ccfDNA Tubes was measured at different storage intervals. Shown are the relative signals of the 3 mutations covered by the therascreen EGFR assay: [A] point mutation T790M, [B] deletion in exon 19 and [C] L858R. Each PCR was done in duplicate. To calculate the delta-C_T value, the average C_T of each mutant was subtracted from the average C_T of the wildtype sub-assay. The average of all donors is shown in the figure for all tested time points. The therascreen EGFR assay defines a threshold for each sub-assays. Delta-C_T values above this threshold result in false-negative calls. Results from the ccfDNA isolated from EDTA tubes exceeded this threshold over time whereas results from the PAXgene Blood ccfDNA Tubes remained stable.

Two protocol lines of the QIAsymphony PAXgene Blood ccfDNA Kit for different research needs.

With the standard protocols (STA), predominantly small ccfDNA fragments are isolated. The large fragment protocols (LAF) efficiently co-purify large and small ccfDNA fragments. Both protocol lines consist of one protocol for 2.4 ml and one protocol for 4.8 plasma input.

The QIAsymphony SP processes samples containing magnetic particles.

A magnetic rod protected by a rod cover enters a well containing the sample and attracts the magnetic particles. The magnetic rod cover is positioned above another well and the magnetic particles are released. The QIAsymphony SP uses a magnetic head with an array of 24 magnetic rods, and can therefore process up to 24 samples simultaneously. Collection, transfer and release of magnetic particles is repeated several times during sample processing.

Isolated ccfDNA is compatible with methylation-specific downstream applications.

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).

Performance

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.

Principle

Apoptosis and lysis of white blood cells in unstabilized whole blood releases genomic DNA into plasma, which dilutes naturally occurring ccfDNA. PAXgene Blood ccfDNA Tubes contain an additive that prevents blood coagulation and immediately stabilizes blood cells via a non-crosslinking stabilization solution (see figure PCR results from blood collected in PAXgene Blood ccfDNA Tubes remain stable over storage time). This impedes the release of intracellular DNA and maintains constant ccfDNA levels in plasma. When used with PAXgene Blood ccfDNA Tubes, the proven magnetic particle technology of the QIAsymphony PAXgene Blood ccfDNA Kit provides a rapid, automated procedure to isolate ccfDNA of high quality and purity (see figure Isolated ccfDNA is compatible with methylation-specific downstream applications).

Procedure

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).

Applications

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:

PCR, digital, multiplex and quantitative real-time PCR
Pharmacogenomic studies
SNP genotyping
Next generation sequencing
Methylation assays

Specifications

Features	Specifications
Applications	Wide spectrum of research applications, including analysis of cancer tumor DNA and non-invasive prenatal testing
Elution volume	60 µl for the STA protocol line 60, 100 or 150 µl for the LAF protocol line
Format	Magnetic beads
Input volume	2.4 ml or 4.8 ml plasma
Processing	Manual: plasma centrifugation Automated: QIAsymphony SP
Shelf life	6 weeks for an open reagent cartridge
Throughput	192 samples per working day and instrument
Time per run	96 samples of 4.8 ml plasma in 5 h 40 min

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FAQs (31)

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	Do I need to use a blood collection set with the PAXgene Blood ccfDNA Tube? FAQ ID - 3626	View
	What is the shelf life of unused PAXgene Blood ccfDNA Tubes? FAQ ID - 3633	View
	Can whole blood in PAXgene Blood ccfDNA Tubes be stored at temperatures below 15°C? FAQ ID - 3641	View
	Can whole blood be frozen in PAXgene Blood ccfDNA Tubes? FAQ ID - 3642	View
	Is double centrifugation required for plasma processing? FAQ ID - 3643	View
	How long and at which temperature can plasma generated from whole blood in PAXgene Blood ccfDNA Tubes be stored? FAQ ID - 3644	View
	How should I thaw frozen plasma samples? FAQ ID - 3645	View
	Which plasma volumes can I process with the QIAsymphony PAXgene Blood ccfDNA protocols? FAQ ID - 3648	View
	How many extractions can I perform with a QIAsymphony PAXgene Blood ccfDNA Kit, using the 2.4 ml or the 4.8 ml protocols? FAQ ID - 3649	View
	How long after first use can a reagent cartridge be stored for further use? FAQ ID - 3650	View
	For what type of sample material is the QIAsymphony PAXgene Blood ccfDNA Kit intended? FAQ ID - 3652	View
	Which tubes are recommended for the QIAsymphony sample rack FAQ ID - 3653	View
	Which protocol line should I use to process plasma samples? FAQ ID - 3654	View
	Does the QIAsymphony PAXgene Blood ccfDNA Kit also allow eluting ccfDNA in less than 60 µl? FAQ ID - 3656	View
	How should eluates produced with the PAXgene Blood ccfDNA System be stored? FAQ ID - 3657	View
	What is the shelf life of the QIAsymphony PAXgene Blood ccfDNA Kit? FAQ ID - 3651	View
	Which downstream applications have been tested with ccfDNA purified from whole blood collected and processed with the PAXgene Blood ccfDNA System? FAQ ID - 3658	View
	Which elution volumes are available with the QIAsymphony PAXgene Blood ccfDNA Kit? FAQ ID - 3655	View
	Why is a blood collection set required to collect blood into the PAXgene Blood ccfDNA Tube? FAQ ID - 3628	View
	Can reagent from the PAXgene Blood ccfDNA Tube flow back into the patient’s arm? FAQ ID - 3629	View
	What is the blood draw volume of the PAXgene Blood ccfDNA Tube FAQ ID - 3631	View
	What is the reagent in the PAXgene Blood ccfDNA Tube? FAQ ID - 3632	View
	How should unused PAXgene Blood ccfDNA Tubes be stored? FAQ ID - 3634	View
	Are there special considerations for phlebotomy when using the PAXgene Blood ccfDNA Tube (blood collection set, discard tube, positioning of the tube, issues with short draws, etc.)? FAQ ID - 3635	View
	Are PAXgene Blood ccfDNA Tubes sterile? FAQ ID - 3636	View
	Can whole blood in PAXgene Blood ccfDNA Tubes be stored at temperatures above 25°C? FAQ ID - 3640	View
	How long can whole blood stabilized in PAXgene Blood ccfDNA Tubes be stored at room temperature? FAQ ID - 3639	View
	Can I use plasma from underfilled PAXgene Blood ccfDNA Tubes? FAQ ID - 3638	View
	Can I use a different brand blood collection set with the PAXgene Blood ccfDNA Tube? FAQ ID - 3630	View
	What is the expected ccfDNA yield from plasma generated from 10 ml of blood? FAQ ID - 3637	View
	What should I do if cryoprecipitates form while thawing a frozen plasma sample? FAQ ID - 3646	View

Scientific Posters (4)

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	A New Blood Collection Tube and Automated Extraction Method for Stabilization and Analysis of ccfDNA Ullius et al., 2015	Show details
	A Shortened, Automated Workflow for Processing Plasma Directly From Primary Blood Collection Tubes Reduces Turnaround Time and Risk of Sample Mix-up in ccfDNA Based Applications Voss et al., 2017	Show details
	Integrated Workflow for Blood Collection, ccfDNA Stabilization and Extraction for Non-invasive Prenatal Testing Ullius et al., 2016	Show details
	Performance Evaluation of a New Integrated Blood Collection and Sample Preparation System for the Stabilization and Extraction of Circulating Cell-free DNA (ccfDNA) Ullius et al.	Show details

Application/Protocol Documents (1)

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	QIAsymphony Protocol Sheet PAXgene ccfDNA_2400 and ccfDNA_4800 For purification of ccfDNA from human whole blood stabilized in PAXgene Blood ccfDNA Tubes using the QIAsymphony SP and QIAsymphony PAXgene Blood ccfDNA Kit.	Show details

Application Notes (1)

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	PAXgene Blood ccfDNA System Primary Tube Handling (EN)	Show details

Brochures & Guides (1)

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	Flyer: Primary Tube Handling with PAXgene Blood ccfDNA System	Show details

Kit Handbooks (1)

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	QIAsymphony PAXgene Blood ccfDNA Kit Handbook For purification of ccfDNA from plasma using the QIAsymphony SP	Show details

Labware Files and Documents (1)

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	Labware List — QIAsymphony PAXgene Blood ccfDNA Kit (SOW-516-7) For use with software version 4.0 or higher	Show details

Safety Data Sheets (1)

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	MSDS QIAsymphony PAXgene Blood ccfDNA Kit (192)

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