Instructions for the laboratory order/pre-analysis

 

1. After having collected the samples order a "Frigobox" by phone (shipping container with dry ice, keeps samples cold up to max. 72h) at +41 71 884 45 45 (Team Labor W AG)
    

2. Upon receipt of the Frigobox, immediately place tubes in the dry ice, place order form on top.

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Collection of the box will be organized by Labor Team W AG.

 

Three different samples should be collected in the following order

(All samples can be drawn using one cannula)

 

1.  Whole blood

 

1. Whole blood should only be collected ONCE during follow-up (at first visit) in K3EDTA tubes. Do not draw and store whole blood more than once if already performed.
    

2. Aliquot (1ml is sufficient) the sample in labeled tubes and immediately freeze in a -70°C to -80°C freezer.No further steps are required.
    

Labeling of samples

<- Every whole blood sample must be labeled according to following scheme. The sticker should specifically be labelled with "WB” to indicate that this aliquot contains whole blood

 

 

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 2.  microRNA

 

1. Collection of approx. 10 ml blood in locally used serum
   tubes (same as for AFP/HCG analyses) and allow sample
   to stand for 30 min at room temperature.
    

2. centrifugation (10 min at 2.500g)
    

3. carefully pipette off the supernatant (serum) immediately,
   transfer to labeled plastic shipping tubes and deep freeze
   at -20°C to -80°C (if the possibility for -80°C is not given,
   it can also be stored temporarily at -20°C for max. 5 days)

 

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Labeling of samples

<- Every sample must be labeled according to following

scheme:

 

In case of positive microRNA-371 test levels, a repeated blood sample will be requested within 4 weeks of the initial measurement.Label the second sample with 1 additional month to the first one, for example first positive test is labeled: “month of follow-up: 03”, and the follow-up sample will be labeled: “month of follow-up: 04”

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3.  Cell free DNA

1. Requirements

1. Use two (2) EDTA tubes (lavender top) to collect whole blood.

2. Use aseptic techniques and draw blood from the patient into the EDTA tube(s). Make sure to collect full volume to ensure correct blood to anticoagulant ratio.

3. Immediately after the blood is drawn, thoroughly mix the blood
   with the anticoagulant by gently inverting the EDTA tube(s) ten (x10) times. 

4. Use aseptic techniques to process whole blood. 

5. Store EDTA tubes at 4°C for a maximum of 2 hours before processing.

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2. Procedure

1. Centrifuge the collection tubes at 2000 x g for 20 minutes at 22º C. Make sure all blood tube processing is performed under a laminar hood to avoid contamination. (If no laminar hood is available, an aseptic working method with sanitized working area and proper personal protective equipment is also possible)

2. Using a sterile transfer pipette, remove the plasma (yellow-clear liquid) stopping
   5mm above the buffy coat (the thin gray-white layer) to ensure no aspiration of
   buffy coat and/or red blood cells/erythrocytesand transfer to a sterile 15 ml centrifuge
   tube. No cells or debris should be present in the plasma. (If no sterile pipettes are
   available, you cansterilize pipettes by autoclaving them or embed on ethanol overnight
   and let dry prior to use)

3. Re-Centrifuge the 15ml centrifuge tube at 3000 x g for 30 minutes at 22ºC and using
   a new sterile transfer pipet, remove the plasma stopping 5mm above any debris pellet. 

4. Aliquot the plasma from each tube into a 1.5mL Eppendorf tubes and label each tube.

5. Immediately freeze the plasma tubes in a -70°C to -80°C freezer.

6. Store frozen plasma tube in a -70°C to -80°C freezer until ready to ship.

 

Labeling of samples:

<- Every sample must be labeled according to following scheme. It is important that this aliquot is labeled as “ctDNA”