Buffy Coat Processing

Acceptable Tube Types
If you plan to extract nucleic acids from materials, Purple-top and Yellow-top tubes are most commonly used.
  • Purple top tube (EDTA anti-coagulant). Commonly 4.0 or 10mL volume tubes for collection of whole blood.
  • Green top tube (lithium heparin anti-coagulant). Commonly 4.0, 5.0 or 10mL tube. Note the heparin anti-coagulant interferes with many nucleic acid extraction protocols
  • Blue top tube (sodium citrate anti-coagulant. Commonly 3.0, 5.0 or 10mL tubes for collection of whole blood. Used primarily for coagulation testing.
  • Yellow top tube (ACD, acid-citrate-dextrose anti-coagulant). Uncommon sample, but often received as 10mL tube.
Standard Procedure
  1. Spin at 3000rpm for 10 minutes at room temperature (25*C).
  2. Use a sterile pipette to pull off plasma, leaving no more than 0.5mL of plasma over the cell interface.
  3. Note the buffy coat/yellowish layer over the packed RBC layer.
  4. With a single-use pipette, collect as much of the yellow layer as possible (generally in <0.5mL volume), avoiding taking significant amounts of plasma or RBCs.
  5. Transfer the buffy coat to a sterile pour-off tube or cryovial.
Variations (added per request to study protocol).
  • Spin time: Can be modified from 10 minutes.
  • Spin temperature: Can be modified from 25*C (4*C-42*C).
  • Storage conditions: ambient, 4*C, -20*C, -80*C.
  • Alternate collection tube, including:
    • Eppendorf tubes: 1.8mL, 2.0mL.
    • Screw capped tubes: 2.0mL, 10mL.
    • Cryovials: specify volume, size (1", 2").