Heart Biomarker Evaluation in Apnea Treatment
4.2.3 Description of Collection Tubes
Tube #1 is a 10mL red-topped Serum tube (B-D 367820). Let these tubes stand at room temperature for a minimum of 30 minutes to allow the blood to clot. They are then centrifuged and the serum is aliquoted (1 clear-capped 4mL tube containing 1.0 mL and 3 red-capped 1.5mL cryovials which contain 1 mL each). The serum will be tested for insulin, glucose and lipid profile as well as stored in repository for later analysis.
Tube #2 is a 5 mL citrate tube (B-D 366415). After centrifugation, the serum is aliquoted (4 blue-capped 0.5 mL cryovials which contain 0.5 mL each). These samples will be used for coagulation testing and stored in repository for later analysis. Once the plasma is removed, the packed cells are removed into one 10 mL red-capped cryovial.
Tube #3 is a 10mL EDTA tube (B-D 366457). After centrifugation, plasma from this tube will be aliquoted (4 purple-capped 1.5 mL cryovials which contain 1 mL each). Once the plasma is removed, the packed cells are removed into the 10 mL red-capped cryovial.
Tube #4 is a 5mL white-topped "Special Coagulation" tube (SCAT-I) containing a white, powdered anticoagulant. This tube contains a special combination of anticoagulants that ensure long-term stability of the plasma sample. Specifically, this tube, when filled, will contain 4.5 mM EDTA, 150 KIU/mL aprotinin and 20 uM D-Phe-Pro-Arg-chloromethylketone. The SCAT-1 tube must be stored refrigerated. It must be drawn after at least one other tube has been drawn. Important to note, this tube is 'non-sterile', therefore it must be drawn using a butterfly apparatus with 12 inches of tubing; alternatively, a syringe may be used for the venipuncture, and expressed through the cap (with great care to limit turbulence) into the SCAT tube. It is critical that the SCAT-1 tube is mixed well (>30 sec of gentle inversion) before being placed on ice to await further processing/centrifugation. The plasma will be aliquoted after centrifugation (4 yellow-capped 0.5 mL cryovials with 0.5 mL each). The plasma will be used for measuring inflammatory markers. Once the plasma is removed, the packed cells are removed into the 10 mL red-capped cryovial.
Tube #5 is an 12 x 125mm, 8mL cell preparation (Cell Prep Tubes or CPT) tubes with green/red (Becton Dickinson # 362753 (heparin)). This tube contains anticoagulant as a polyester gel and a density gradient liquid (Ficoll Hypaque) to enhance the separation of white cells. These tubes will be processed to obtain viable white cells (a “buffy coat” layer appears in the density gradient). After filling with blood, these tubes are mixed (either by gentle inversion or on a mixer) and kept at room temperature. Tubes must be processed within two hours of collection.