What are the precautions that should be considered in collecting blood for coagulation studies from an indwelling catheter?

Objectives: To determine the amount of blood that should be discarded from a peripheral normal saline lock, a capped-off intravenous port, before a blood sample is obtained for determination of activated partial thromboplastin time from patients being treated with heparin.

Methods: A prospective, quasi-experimental design was used with 32 patients. A blood sample was obtained via venipuncture from each patient to serve as the control for that patient. The normal saline lock was flushed with 2 mL of normal saline. Four consecutive 3-mL blood samples were obtained directly from the normal saline lock, representing samples obtained after discard volumes of 0, 2, 4, and 6 times the dead space of the catheter and extension set (1.5 mL). Activated partial thromboplastin times for the venipuncture blood sample were compared with the times for the blood samples obtained from the normal saline lock.

Results: The only significant difference (P = .02) was that activated partial thromboplastin time was 15% higher in the blood sample obtained from the normal saline lock with no blood discarded than in the venipuncture blood sample.

Conclusions: Nurses can obtain accurate measurements of activated partial thromboplastin time with blood samples obtained from normal saline locks by first discarding a volume of blood equal to 2 times the dead space of the catheter and extension set. Obtaining blood samples in this manner reduces patients' discomfort due to repeated venipuncture and diminishes blood loss.

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Specimen Transportation

	PRC-PALM-SPC-5.51-pro1-prs3: COAGULATION SPECIMEN COLLECTION AND TRANSPORT Specimen Collection Collect specimen in blue top (3.2% sodium citrate) tube. If multiple tests are being drawn, draw coagulation studies before additive-containing tubes such as the EDTA, heparin, or clot activator (SST) tubes. If the coagulation tube is being drawn using a winged collection device with variable tubing length so air in the tubing is not introduced into the blood collection tubes leading to under-filling, draw 1-2 mL into another blue top (3.2% sodium citrate) Vacutainer®, discard, and then collect the specimen. When using the hypodermic needle/syringe, it is important the blood is added to the appropriate volume of anticoagulant within one minute of completion of draw. Once obtained, the tube should be gently inverted three or six times end over end inversions; do not over mix as excessive mixing will affect the test result. Insufficient mixing may have a greater effect on specialized hemostasis assays. Citrate tubes must be adequately filled (to the mark noted on the tube if provided) or to not less than 90 % of this total volume. Inadequate filling of the collection tube will lead to inaccurate results. The final citrate concentration in the blood should be adjusted in patients who have hematocrit values above 0.55L/L (55%). For hematocrits below 02.0L/L (20%), there are no current data available to support a recommendation for adjusting the citrate concentration. PRC-HFH-COA-5.51-stw3: VISUAL AID OF VOLUME REQUIREMNTS FOR COAGULATION SAMPLES STANDARD WORK When drawing the specimen, avoid contaminating the sample with tissue thromboplastin as this may affect results. Venipuncture must be clean with no trauma, and the application of the tourniquet should be limited to 1 minute. Collection of the blood through lines that have been previously flushed with heparin should be avoided. If the blood must be drawn through a VAD (vascular access device), possible heparin contamination and specimen dilution should be considered.  In this case the line should be flushed with 5 mL of saline and the first 5 mL of blood or six dead space volumes of the VAD discarded. Blood should never be transferred from 1 collection tube to another in effort to provide the required fill volume. This is true even if two sodium citrate tubes are combined as this may lead to doubling up of anticoagulant citrate levels and further dilution of the plasma sample.   Transport of Whole Blood Specimens Store and transport whole blood specimens at room temperature. Avoid exposure of whole blood samples to ice or freezing temperatures. Unless otherwise directed, transport the specimen to the laboratory at room temperature within two hours of collection. PT specimens from the Medical Centers may be transported to the laboratory at room temperature for up to 24 hours after collection. APTT from the HFHS Medical Centers:If patient is not receiving heparin, specimen may be sent in the native tube at room temperature or in a common cooler for up to 12 hours after collection. If delivery to the laboratory will be delayed more than 12 hours, centrifuge specimen at 1500xg (at least 3000 rpm) for 15 minutes or at a centrifuge speed to consistently produce platelet poor plasma. Transfer platelet poor plasma to a polypropylene tube and cap. Avoid contamination of plasma with platelets from the buffy coat layer. Freeze plasma rapidly. Transport specimen to laboratory on dry ice in a suitable container. Specimens for other assays (e.g. thrombin time, protein C, Factor V and other Factors) kept at 2°- 4°C or 18 to 24°, should be centrifuged and tested within four hours from time of specimen collection. Specimens for tests other than PT/APTT that are collected at Medical centers will require special processing (please refer to the specific tests page for additional instructions as needed). Centrifuge the tube at 1500 x g (3000 RPM) for 15 minutes to separate the plasma. Transfer the platelet poor plasma to a 12x75 mm polypropylene tube. Avoid contamination of plasma with platelets from the buffy layer. Securely cap the tube. Properly label the tube and freeze rapidly at -20°C or below. Specimens can be frozen up to two weeks at -20° or for six months at -70°. Transport specimen to the laboratory on dry ice in a suitable container. DO NOT POOL SPECIMENS FOR COAGULATION. EACH TUBE SHOULD BE ALIQUOTED INTO AN INDEPENDENT TUBE PRIOR TO FREEZING. The number of tests that can be done depends on the combination of tests requested, condition of the sample(s), and potential need for reflexive testing. Specimens that are improperly filled (under filled/QNS or overfilled), clotted, hemolyzed, received after prolonged delay, collected in an improper container (wrong tube/anticoagulant), collected with the wrong sodium citrate concentration, or collected in an expired collection tube will be rejected. For technical information, contact the Special Coagulation Laboratory at (313) 916-1825.   Last Modified: Friday, March 25, 2022 11:05 AM
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1. Collection Tube. Blood should be collected in a blue-top tube containing 3.2% buffered sodium citrate.

Note: Please examine specimen collection and transportation supplies to be sure they do not include expired containers.

2. High Hematocrit Samples. Patients with elevated hematocrits have a relatively low amount of plasma for a given whole blood (collection) volume. This tends to increase the plasma citrate concentration effectively. If the patient has a known hematocrit >55%, the amount of citrate in the collection tube must be decreased according to the formula below.

The equation in CLSI H21 is as follows:

C = (1.85 x 10−3) (100−Hct) (Vblood)

Where:

C is the volume of citrate remaining in the tube.

Hct is the patient's hematocrit.

V is the volume of blood added to the evacuated tube.

Example:

Patient hematocrit = 60%

Total volume = 5 mL (standard citrated plasma collection tube volume)

C = (0.00185) x (100-60) x 4.5

C = (0.00185) x 180 (or 40 x 4.5)

C = 0.333

3. Order of Draw. A discard tube is not required prior to collection of coagulation samples, except when using a safety winged blood collection device (ie, "butterfly"), in which case a discard tube should be used. When noncitrate tubes are collected for other tests, collect sterile and nonadditive (red-top) tubes prior to citrate (blue-top) tubes. Any tube containing an alternate anticoagulant should be collected after the blue-top tube. Gel-barrier tubes and serum tubes with clot initiators should also be collected after the citrate tubes.

4. Venipuncture Technique. To avoid activation of the sample, the venipuncture should be clean, with minimal trauma. The tourniquet should be in place only as long as is needed to identify a vein and should never be in place for longer than one minute. Severely hemolyzed samples are not acceptable.

5. Safety winged blood collection kits (butterfly) must use a discard lead tube prior to collecting specimen tube to submit for testing. Failure to use a discard tube may lead to underfilling of the evacuated tube.

6. Fill Volume. Evacuated collection tubes must be filled to completion to ensure that a 9:1 blood-to-anticoagulant ratio is achieved. Under-filling of citrate collection tubes results in an increased anticoagulant-to-blood ratio and can extend clot-based coagulation assays. Note: Never combine two underfilled tubes together.

Please print and use the Volume Guide for Coagulation Testing to ensure proper draw volume.

7. Mixing. The sample should be mixed immediately by three to six complete gentle end-over-end inversions to ensure adequate mixing of the anticoagulant with the blood.

8. Plasma Processing. Process the sample as soon as possible (preferably within 30 minutes of collection). Centrifuge at an adequate speed and duration to achieve platelet-poor plasma (<10,000/μL). After centrifuging the sample, transfer plasma, being careful not to approach the buffy coat, using a plastic pipette into a LabCorp PP transpak frozen purple tube with screw cap (LabCorp N° 49482). Note that glass should not be used because glass can activate the clotting cascade. Label each tube “plasma, citrate.” The specimen should be frozen immediately and maintained frozen until tested. To avoid delays in turnaround time when requesting multiple tests on frozen samples, please submit separate frozen specimens for each test requested.

Platelet-poor Plasma (PPP) Collection for Lupus Anticoagulant Testing

Lupus anticoagulants (LA) are are nonspecific antibodies that extend clot-based coagulation assays as the result of their interaction with phospholipid in the reaction mixture. Platelets in plasma samples can act as a source of phospholipid and mask the effects of LA. For this reason, it is important to prepare platelet-poor plasma (PPP) for LA testing. PPP should have a platelet count <10,000/μL. PPP samples should be collected by double centrifugation.

1. Centrifuge for 10 minutes, and carefully remove two-thirds of the plasma using a plastic transfer pipette, being careful not to disturb the cells.

2. Deliver plasma to a plastic transport tube, cap, and recentrifuge for 10 minutes.

3. Use a second plastic pipette to remove the plasma, staying clear of the platelets at the bottom of the tube.

4. Transfer plasma using a plastic pipette into a LabCorp PP transpak frozen purple tube with screw cap (LabCorp N° 49482). Label each tube "plasma, citrate."

The specimen should be frozen immediately and maintained frozen until tested. To avoid delays in turnaround time when requesting multiple tests on frozen samples, please submit separate frozen specimens for each test requested.

Clearing (Flushing) the Volume of Intravenous Lines Before Drawing Samples for Hemostasis Testing

Collection of blood for coagulation testing through intravenous lines that have been previously flushed with heparin should be avoided, if possible. If the blood must be drawn through an indwelling catheter, possible heparin contamination and specimen dilution should be considered.

When obtaining specimens from indwelling lines that may contain heparin, the line should be flushed with 5 mL of saline, and the first 5 mL of blood or six times the line volume (dead space volume of the catheter) should be drawn off and discarded before the coagulation tube is filled. For those samples collected from a normal saline lock (capped off venous port), twice the dead space volume of the catheter and extension set should be discarded.

References

Adcock DM, Kressin DC, Marlar RA. Are discard tubes necessary in coagulation studies? Lab Med.1997; 28:530-533.

Brigden ML, Graydon C, McLeod B, Lesperance M. Prothrombin time determination. The lack of need for a discard tube and 24-hour stability. Am J Clin Pathol. 1997 Oct; 108(4):422-426.

Clinical and Laboratory Standards Institute. Collection, Transport, and Processing of Blood Specimens for Testing Plasma-Based Coagulation Assays; Approved Guideline—Fifth Edition. Wayne, Pa: CLSI; 2008.

Konopad E, Grace M, Johnston R, Noseworthy T, Shustack A. Comparison of PT and aPTT values drawn by venipuncture and arterial line using three discard volumes. Am J Crit Care. 1992 Nov; 1(3):94-101.

Laxson CJ, Titler MG. Drawing coagulation studies from arterial lines; an integrative literature review. Am J Critical Care. 1994 Jan; 3(1):16-24.

Lew JK, Hutchinson R, Lin ES. Intra-arterial blood sampling for clotting studies. Effects of heparin contamination. Anaesthesia. 1991 Sep; 46(9):719-721.