Processing Off-site Coagulation Specimens

The following procedural guidelines are for normal coagulation specimens collected at the off-site facilities and transported to the main laboratory for testing. The following guidelines may be used for non-therapeutic Protimes [PT], therapeutic Protimes [TPT]- patient on Coumadin therapy, non-therapeutic APTTs, D-Dimers, and Fibrinogens.

Patients who are receiving Heparin therapy MUST have special arrangements made for the collection and transport of their specimens.

Contact Coagulation Department (583.6797) prior to drawing TAPTT or regarding any “special” coagulation tests [e.g. Lupus Anticoagulants, Protein C, Protein S, Factor assays]

Specimen Requirements

  1. Refer to the Basic Venipuncture Procedure, Blood Collection Vascular Access Devices, and/or Fingerstick Skin Puncture Procedure, for specific collection guidelines.
  2. Coagulation tests other than PT, APTT, TPT, and TAPTT, should not be collected first, a discard tube (blue top) should be collected first, to prevent any contamination of specimen with “tissue thromboplastin, which will alter test results.
  3. PROMPT AND ADEQUATE MIXING IS ABSOLUTELY ESSENTIAL .
  4. Greiner Bio-One VACUETTE® plastic vacutainer tube with a light blue cap is the collection tube of choice:
    • Contains 3.2% Sodium Citrate
    • Invert 4 times after filling
    • Draw volume is 2.7 mls
    • Must be filled to the fill line printed on the tube. The evacuated tube vacuum will fill the tube to the fill line. Collection volume below 90% is unacceptable and the specimen will be rejected.
    • The ability for the tube to prevent the blood from clotting is dependant on the concentration of anticoagulant vs. the volume of blood collected. The hematocrit of the patient is also a factor when assessing the proper concentration of anticoagulant. If a patient’s hematocrit appears to be greater than 55%, contact the Coagulation Department (583.6797) for information on adjusting the amount of citrate needed to collect the blood specimen. This adjustment may be needed specifically for pediatric-neonate patients or polycythemic patients.

      Since the main lab receives the processed plasma specimens, it is imperative that the person processing the specimen observes for the presence of an elevated RBC count. If the centrifuged specimen has over 55% red cell layer, please notify the Hematology department immediately.

Centrifugation

  1. Most of the plasma coagulation tests are performed on platelet-poor plasma (less than 10,000/ cumm) obtained by centrifugation at 1600-2000 g (3000 rpm) for 10-15 minutes or by centrifugation at 4440 g (7200 rpm) for 3 minutes.
  2. Specimens are to remain capped through centrifugation, processing (as much as possible), storage, and transport.
  3. Following centrifugation, filter the cell-free plasma from the red cells and buffy coat using a S/P Brand Filter, and aliquot into a blue polypropylene micro-centrifuge tube(s) or 12x75 cloudy polypropylene tubes and seal tightly.
  4. If more than just a PT/APTT is requested, process several aliquots of the specimen. [For “Special” coagulation tests contact the Coagulation Department (583.6797) for instructions.]
  5. Proper labeling of specimen is essential, and MUST include collection time and initials.

Specimen Appearance

  1. Check specimen for:
  2. Visible appearance of a partial clot
    • Icteria (yellow tinged plasma)
    • Hemolysis
    • Cancel and request recollection of all hemolyzed or clotted specimens; document in the computer person called, date, and time.
  3. Collection volume must be within 90% of required amount for the collection tube. Cancel and reorder any “short draw” specimens, document in the computer person called, date, and time.

    ALL SPECIMENS ARE POTENTIALITY HAZARDOUS AND MUST BE HANDLED FOLLOWING UNIVERSAL PRECAUTION PROCEDURES.

Specimen Freezing and Storage

  1. After processing specimen, place labeled aliquot in freezer in the supplied freezer specimen container. Specimens are to be frozen [minimum -20 ° C] immediately after collection and remain frozen throughout storage and transportation to the main laboratory.
  2. Specimens for TAPTT cannot be frozen . Call Coagulation Department (583. 6797) for instructions prior to collection.
  3. Each off-site facility is to be supplied with three freezer containers [DyNACHILL] that are kept in the freezers and rotated to and from the main laboratory by the couriers. The freezer containers DyNACHILL will be labeled with the name of the collection site and are to be returned to that site. Some couriers carry dry ice. In these cases a DyNACHILL container is not needed. Care must be taken that the specimens do not come in direct contact with the dry ice, the tubes may crack in a rare situation.
  4. Upon return to the off-site facility, the opened freezer container DyNACHILL, including lid, must be placed in the freezer for a minimum of 4 hours, {if not overnight} before used again for processing specimens.
  5. Each off-site facility will be responsible for disinfecting the freezer containers, both internally and externally, with an approved disinfectant, at their location.

Specimen Transportation

  1. Couriers will pick up the DyNACHILLs from the off-site facility and deliver them to the Main Lab Business Office for further processing.
  2. Office staff will deliver the specimens and specimen labels [leaving the specimens in the DyNACHILL] to the Coagulation Department for testing.
  3. All specimens are to be received at the Covenant HealthCare Main Laboratory; on the day of collection, by 8 PM for analysis. Specimens are NOT to be kept over night at the off-site facilities.
  4. Testing will take place within ½ hr. after the thawing of the specimens.
  5. The DyNACHILLs will be returned to the courier pickup room, to be returned to the off-site facilities.

Specimen Stability

Specimen stability is affected by:

  1. Preservation of Factors V and VIII, processing in opened tubes means a loss of dissolved CO 2 and subsequent pH change.
  2. As platelets break down and release their coagulation factors many changes may take place. Specimens are to be centrifuged and filtered to remove all cellular material prior to freezing of plasma.
  3. Glass contact; plastic test tubes must be used for specimen storage.
  4. Choice and concentration of anticoagulant and its buffering capacity.
  5. Temperature and time of storage.
  6. Size and shape of collection tube.
  7. Handling of specimen during processing.
  8. Specimens frozen in polystyrene tubes are not acceptable. They activate the clotting system.

References

  1. National Committee for Clinical Laboratory Standards, NCCLS Publication: Vol. 2 NO. 4, “Tentative Guidelines for the Standardized Collection, Transport and Preparation of Blood Specimens for Coagulation Assays. Pub. April 1982
  2. “Blood Coagulation” Clinical and Laboratory Aspects” , University of Michigan, 1979
  3. “Hemostasis and Thrombosis” Robert W. Colman MD., Jack Hirsh MD., Victor Marder MD., Edwin Salzman MD., Second Edition 1987
  4. “Hemostasis and Thrombosis in the Clinical Laboratory”, Donna M. Corriveau, M.A., M.T.(A.S.C.P.), George A. Fritsma, M.S. M.T.(A.S.C.P.) J. B. Lippincott Co. 1988
  5. NCCLS publication : “CLINICAL LABORATORY PROCEDURE MANUALS”, NCCLS VOL.4 NO. 2 College of American Pathologists, and Commission on Laboratory Accreditation, CAP 1991.

 

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