Summary and Principles 

CelliMAX Cell Free DNA Tube stabilizes cell-free plasma DNA as well as preserves cellular genomic DNA present in nucleated blood cells and circulating epithelial cells (tumor cells) found in whole blood. Accurate analysis of cf-DNA can be compromised by sample handling, shipping and processing, caus in glys is of nucleated blood cells and subsequent release of cellular genomic DNA. Additionally, degradation of cf-DNA due to nuclease activity can be problematic. The preservative reagent contained in CelliMAX Cell Free DNA Tubes tab ilizes nucleated blood cells, preventing the release of cellular genomic DNA, and inhibits nuclease mediated degradation of cf- DNA, contributing to the overall stabilization of cf-DNA. Samples collected in CelliMAX Cell Free DNA Tube are stable for up to 14 days at temperatures between 6°C to 37°C, allowing convenient sample collection, transport and storage. The preservative reagent present in CelliMAX Cell Free DNA Tube Stabilizer Circulating Epithelial cells (tumor cells) in whole blood for up to 7 days at temperatures between 15°C to 30°C.

Reagents

CelliMAX Cell Free DNA Tube contains the anticoagulant K3 EDTA and a cell preservative in a liquid medium.

Indications of Product Deterioration

1. Cloudiness or precipitate visible in reagent of empty tube.

2. If indications of product deterioration occur, contact us

Instructions For Use

For a video demonstration, visit MP Biomedical.com/ mixing.

1. Collect specimen by venipuncture according to CLSI Gp411. Prevention of Back flow-Since CelliMAX Cell Free DNA Tube contains chemical additives, it is important to avoid possible back flow from the tube. To guard against back flow, observe the following precautions:

a.  Keep patient's arm in the down ward position during the collection procedure.

b.  Hold the tube with the stopper in the uppermost position so that the tube contents do not touch the stopper or the end of the needle during sample collection.

c.  Releaset our niquet once blood starts to flow in the tube, or with in 2 minutes of application.

•   Follow recommendations for order of draw outlined in CLSI Gp411.CelliMAX Cell Free DNA Tube should be drawn after the EDTA tube and before fluoride oxalate (glycolytic inhibitor) tube. If a CelliMAX Cell Free DNA Tube tube immediately follow sahepar in tube in the draw order, MP Biomedical recommends collecting a non-additive or EDTA tube as a waste tube prior to collection in the CelliMAX Cell Free DNA Tube.

•   Fill tube completely.

•   Remove tube from adapter and immediately mix by gentle in version 8 to 10 times. In adequate or delayed mixing may result in incorrect analytical results or poor product performance. One in version is a complete turn of the wrist, 180 degrees, and back per the figure below:

2.  After collection, transport and store tubes with in the recommended temperature range.

3.  When using a winged (butterfly) collection set for veni puncture and the MP Biomedical CelliMAX Cell Free DNA Tube is the first tube drawn, a non-additive or EDTA discard tube should be partially drawn first in order to eliminate air or “dead space” from the tubing

4. CelliMAX Cell Free DNA Tube does not dilute blood samples; there fore, no dilution fac tor correction is necessary.

5. As in the case with most clinical laboratory specimens, hemolysis, icterus and lipemia may affect the results obtained on blood samples preserved with CelliMAX Cell Free DNA Tube.

DNA Extraction 

Cell-Free Plasma DNA 

MP Biomedical has qualified two separate plasma separation spin protocols for your convenience.

Double Spin Protocol

Step 1. To separate plasma, centrifuge whole blood at 300 x g for 20 minutes at room temperature.

Step 2. Remove the upper plasma layer and transfer to a new conical tube (not provided).

Step 3. Centrifuge the plasma at 5000 x g for 10 minutes.

Step 4. Isolate cell-free DNA per kit manufacturer instructions.

Double Spin Protocol (for maximum plasm a recovery) 

Step 1. To separate plasma, centrifuge whole blood at 1600 xg for 10 minutes at room temperature.

Step 2. Remove the upper plasma layerand transfer to a new conical tube (not provided).

Step 3. Centrifuge the plasma at 16000 x g for 10 minutes.

Step 4. Isolate cell-free DNAperkit manufacturer instructions.

For optimal results, include a Proteinase K treatment step (>30 mAU/ mL digest) at 60°C in the presence of chaotropic salts for 1 hour when extractingcell-free DNA.

Cellular Genomic DNA 

Step 1. To separate the white blood cells, either lyse the red blood cells and wash, or centrifuge whole blood and collect the buffy coat layer.

Step 2. Isolate genomic DNA per kit manufacturer instructions

For optimal results, include a Proteinase K treatment step(>30m

AU/mldigest) at 60°C in the presence of chaotropic salts for 2 hours when extracting cellular genomic DNA.

Storage and stability

1.  When stored at 2°C to 30°C, empty CelliMAX Cell Free DNA Tube is stable through expiration date.

2.  Short term storage at 2°C to 40°C is acceptable for empty CelliMAX Cell Free DNA Tube for up to14 days.

3.  Do not freeze empty CelliMAX Cell Free DNA Tube. Proper in sulati'on may be required for shipment during extreme temperature conditions.

4.  Sample storage/stability:

Freezing and Thawing

Plasma

1. To Freeze: Forlong-term storage, after spinning, collect and transfer the upper plasma layer to acryogenic tube (not provided) and freeze at -20 °C or -80 °C.

2. To Thaw: Thaw cryogenic tubes at appropriate temperature as specified in your protocol. Note: If cryoprecipitates form in the plasma, vortex the tube for 30 seconds after thawing. Do not centrifuge the plasma.

Limitations

1.  For single use only.

2. Samples drawing the ranticoagulants or preservatives may cause coagulation in CelliMAX Cell Free DNA Tube.

3. Specimen transport vial pneumatic tube system is not advised.

Precautions

1. For Research Use Only. Not for use in diagnostic procedures.

2. Do not frozen specimens collected in glass CelliMAX Cell Free DNA Tube.

3. Do not use tubes after expiration date.

4. Do not use tubes for collection of materials to be injected in to patients.

5. Product is intended for use as supplied. Do not dilute pradd other components to CelliMAX Cell Free DNA Tube.

6. Over fill in go under filling of tubes will result in an incorrect blood- to-add live ratio and may lead to in correct analytic results or poor product performance.

Caution

a. Glass has the potential for breakage; precautionary measures should be taken during handling of glass tubes.

b. All biological specimens and materials coming in contact with them are considered biohazards and should be treated as if capable of transmitt'ng infection. Dispose of in accordance with Federal, state and local regulations. Avoid contact with skin and mucous membranes.

c.  Product should be disposed with infecti'onus medical waste.

d.  Remove and reinsert stopper by either gently rocking the stopper from side to side or by grasping with a simultaneous twisting and pulling action. A “thumb roll” procedure for stopper removal is NOT recommended as tube breakage and injury may result.

SD Scan be obtained at www.mpbio.com 

References

1. Clinical and Laboratory Standards Institute. Gp41, Procedures for the collection of diagnostic blood specimens by venipuncture. Approved Standard-Seventh Edition.