Coagulation testing is highly sensitive to a myriad of specimen collection and handling variables. Because important diagnostic and therapeutic decisions are based on the results of coagulation tests, a procedural guideline is required to fully address these variables. The following information applies to all Routine Coagulation tests (i.e., Prothrombin Time, APTT, Fibrinogen, etc.). For details on Special Coagulation tests (Lupus Panel, DRVVT, Platelet Aggregation, etc.) refer to the individual test procedure or the Laboratory Survival Guide.
Hemoguard Closure tubes, 4.5mL: This is the "preferred" tube for most Coagulation testing. It is a plastic tube that is identical to the conventional glass or plastic 4.5 mL vacutainer tubes, with the added safety feature of a protective plastic cover over the rubber stopper. These tubes can be placed directly on Coagulation instrument after centrifugation.
Conventional Closure tubes, 4.5 mL: Conventional closure glass vacutainer tubes are still in circulation, but are not the recommended tube for Coagulation testing.
Conventional Closure Pediatric tubes, 2.7mL, 1.8 mL: Conventional closure glass vacutainer tubes are available in pediatric volume sizes.
INDWELLING CATHETER COLLECTION
CLSI RECOMMENDED ORDER OF DRAW:
By following the following "order of draw," many pre-analytical errors and cross-contamination of additives will be avoided.
UNDERFILLING: Inadequate filling of the collection tube will decrease the required blood:anticoagulant ratio (9:1), and may lead to falsely prolonged results.
OVERFILLING: Overfilling will cause an incorrect blood to anticoagulant ratio which may result in slightly decreased results or the specimen may clot or contain fibrin due to inadequate anticoagulant for the blood volume drawn which will result in the specimen being rejected.
HIGH HEMATOCRIT: Results on patients with a high hematocrit (³55%) may be falsely prolonged. Refer to the section (below) on special handling of specimens with HIGH HEMATOCRITS.
CLOTTED SPECIMENS: Clotting will lead to erroneous results. CLSI recommends 5-10 inversions to prevent clotting. Each tube is visually examined prior to centrifugation for the presence of obvious clots. If in doubt, the specimen is checked for clots using applicator sticks.
HEMOLYSIS: Hemolysis of the RBC's causes release of hemoglobin into the plasma, which shortens the APTT test. Hemolysis also suggests the possibility of in vitro clotting, which would adversely affect ALL coagulation tests. Therefore, samples that have visible hemolysis are rejected and a redraw requested. If the level of hemolysis is in question, the specimen will be processed using a “Downtime Label” to determine acceptability.
SPECIAL COAGULATION TESTS: CLSI guidelines recommends drawing a discard tube before specimens collected for special coagulation tests.
CONDITIONS of TRANSPORT and STORAGE
Coagulation tests are enzymatic procedures and, as such, are subject to stringent time-frame and storage guidelines. Reaction temperatures and the pH of specimens must be controlled at all times. For best results, most sources recommend that specimens for Coagulation testing be delivered to the laboratory for testing within 1/2 hour of collection. Considering the logistics and problems associated with transporting specimens, this recommendation is rarely achieved. More realistic time-frame guidelines have been established and must be adhered to. Receipt in lab beyond the stated guidelines will result in REJECTION of the specimen. The allowable time interval between collection of the specimen and testing of the sample will depend on the transport temperature and the storage of the specimen. Specimens for coagulation testing should be processed/stored as follows:
Most specimens for routine Coagulation testing can be transported either as whole blood or centrifuged (plasma) form. If plasma is sent, proper centrifugation protocol must be followed.
Specimens for routine Coagulation testing should be transported either at room temperature* (18-24°C) or refrigerated (2-4°C). *Specimens for Prothrombin Time testing (PT) should be transported at room temperature. They should NOT be refrigerated.
Snap freezing platelet-free plasma (-70°C) is acceptable ONLY if performed by trained technical personnel. There are inherent problems associated with proper freezing/ thawing techniques that can result in inaccurate test results. If properly frozen (-70°C) and aliquotted (capped plastic cryo-tubes), plasma is viable for Coagulation testing for 6 months or at -20°C for 2 weeks.
All specimens received in this laboratory must be properly identified and be accompanied by an appropriate requisition or manifest. Laboratory personnel are required to stringently check identification and test request information prior to processing specimens. Results will NOT be reported on any patient specimen if there is any doubt as to the validity of the requisition and/or the identity of the specimen.
SPECIMEN QUANTITY vs TUBE SIZE
CHECKING FOR CLOTS
After identification and test request is confirmed, volume is checked, and the specimen is checked for presence of clots, the tube is labeled with a Laboratory Information System (LIS) label. LIS label information is checked against requisition slip and patient identification information prior to testing.
For electromagnetic viscosity detection instruments (i.e., STAR, Compact), fast-spun platelet-poor plasma is preferred ( < 10,000/ul). Routine Coagulation testing in this laboratory utilizes the STAT60 centrifuge to obtain this plasma. Quality Control checks on platelet counts are performed on a semi-annual basis, and documented in the Centrifuge Maintenance log.
Note: always invert specimen 2-3 times before centrifugation to facilitate achieving low residual platelet counts.
SPECIAL COAGULATION TESTS
Special Coagulation tests or frozen plasmas require double spun Platelet-Free plasmas.
Special tests that require double spun to obtain platelet-free plasma are:
After centrifugation, specimens are loaded onto the appropriate Coagulation Instrument (Stago: STAR or COMPACT) for Routine Coagulation testing.
After all tests are run, the original collection tubes (and/or remaining transferred plasma) are refrigerated and retained (capped), for 24 hours. Tubes are stored according to shift of testing.
If Special Coagulation tests are ordered in addition to the Routine test, laboratory personnel will refer to special handling instructions included in the individual test procedure.
After 24 hours, specimens are disposed of in specially designated RED BIOHAZARD BAGS/BOXES.
It is the
responsibility of the laboratory to detect any specimens received for
coagulation testing that might have an elevated hematocrit.
Specimens with extremely elevated hematocrits (³55%)
require special handling for Coagulation testing due to improper plasma
to anticoagulant ratio.
When a specimen with a hematocrit ³55% is identified, the laboratory will notify the attending RN or MD and prepare special tubes to be used for coagulation testing.
The lab has two mechanisms to detect specimens with high hematocrits:
If a patient specimen is determined to have a hematocrit ³ 55%, a special tube is prepared by laboratory personnel to adjust the final citrate concentration of the specimen. The method for tube preparation is cited below. The goal is to achieve a blood : anticoagulant ratio of 9:1.
Lab personnel will provide the tube(s) for collection.
FORMULA: y = amount of whole blood required per 0.25 ml Sodium Citrate
y = [( 60 ) X 4.5 ] ¸ 2
100 - Hematocrit
Preparation of "special" tubes for polycythemic patients (HCT ≥ 55%):
NOTE: This can be accomplished by checking the LIS for concurrent CBC results, running original blue-top (citrate tube) on CBC analyzer [multiply result by 1.1 to compensate for blood:anticoagulant difference], or requesting that a new CBC be drawn. One of the first two options is preferable.