Laboratory Methods of Heparin Monitoring, Part 2

College of American Pathologists on unfractionated heparin
UFH is commonly administered as a continuous intravenous (IV) infusion or subcutaneous (SC) injection. The current standard on initial dosing is weight-based and dependent upon the patient’s indication; whether it is for the treatment of venous thromboembolism, coronary thrombosis syndrome, or myocardial infarction.18 Because UFH exhibits inter-individual variability and intra-individual instability, laboratory monitoring is warranted.19

The College of American Pathologists (CAP) has published recommendations regarding laboratory monitoring of UFH.20 Highlights of the general recommendations are as follows: (1) therapeutic heparin monitoring requires the use of a method with a defined therapeutic range; (2) upon UFH infusion or after dose adjustment, anticoagulant monitoring should be done at 6-hour intervals until a stable therapeutic response is reached; then, daily thereafter; (3) daily sampling time should be standardized preferably prior to 10 AM; (4) specimens should be collected by venipuncture from an extremity other than the one used for heparin infusion to avoid possible contamination; (5) clinicians should be informed of the method used to monitor heparin therapy.4,20

Activated partial thromboplastin time – the “Old Standard”
Activated partial thromboplastin time (APTT) is the most widely used test to monitor unfractionated heparin.3,5-8,10,18,20 If this is the method of choice, then only the most accurate protocol for determining the heparin therapeutic range should be used. The CAP recommends that each laboratory develop their own therapeutic range for their APTT system by comparing ex vivo specimens with an appropriately validated heparin assay(ie protamine sulfate neutralization or anti-factor Xa).20 The historic and widely used practice of heparin anticoagulation that is 1.5 to 2.5 times the local APTT control is antiquated and an unsafe practice which could lead to subtherapeutic patient management.17,20 Furthermore, it is also recommended that the therapeutic range be determined with each change in reagent lot number, manufacturer, or instrument. This consensus recommendation illustrates the narrow therapeutic range of UFH. However, the goal of a target therapeutic response still needs to be achieved. This goal is to ascertain the therapeutic value that would reflect maximal antithrombotic effect to prevent recurrence or extension of thromboembolism while maintaining minimal risk of bleeding.2,20

Gausman & Marlar (2013) conducted a study to investigate the optimum number of samples to accurately determine the heparin therapeutic range (HTR) using the ex- vivo method.3 The APTT HTR was calculated with APTT lower and higher limits which correlated to 0.3 U/ml and 0.7 U/ml of heparin, respectively. The study led them to conclude that the optimum number of ex-vivo samples to obtain an accurate HTR determination is 30 samples, but the absolute minimum number is 20. Moreover, acceptable sample requirements for determining the true HTR are: (1) international normalized ratio (INR) should be less than 1.3; (2) less than 50% of the samples have an INR of 1.3 to 1.5; (3) no samples should have an INR greater than 1.5; and (4) less than ten percent of samples are from the same individual.3

The continued use of APTT is due to its numerous advantages: (1) widespread availability; (2) ease and speed of performance; it is easily automated; (3) overall technical reliability; (3) relatively inexpensive cost; and (4) clinicians are generally satisfied with its use.4,8,10,20

However, although APTT is readily used in clinical laboratories, it is noted to have a number of shortcomings that continued to accumulate through the years: (1) poorly standardized; (2) variable patient response; (3) high within and between laboratory variability; (3) biologic variables (heparin binding proteins and factor levels) that can affect monitoring of heparin therapy; (4) pre-analytic variables (underfilling sample tube being the most common); and (5) each laboratory has to determine their own therapeutic range.3,8,9,10,20

The significance of APTT as a monitoring test was assessed in a meta-regression analysis done by Vardi et al (2009). The authors analyzed medical literature for correlation between the level of anticoagulation as measured by APTT to clinical outcomes.11The analysis included 17 publications from 15 randomized clinical trials. A total of 444 and 408 patients with deep venous thrombosis receiving subcutaneous and intravenous UFH, respectively, were included in the study. They used values from the initial phase(defined as the obtained value during the first 48 hours of treatment) and the maintenance phase(defined as the value obtained after 48 hours of treatment, last measure taken, or the mean of the measures obtained). The results were then analyzed using a survey-logistic procedure. The authors concluded that “no correlation between the anticoagulant level and the major clinical outcomes were found, except for the initial anticoagulant measurement and the total mortality at three months, but not to the death related to treatment or disease progression.”

There is still a need for an alternative method that will accurately reflect the anticoagulant effect of UFH in vivo, because of the previously discussed shortcomings of APTT and the test being only a surrogate assay for UFH concentration .

Focus on anti-Factor Xa Assay
Anti- Factor Xa assay, otherwise known as Heparin Xa assay, is currently used as the standard to calibrate the APTT assay, but recently its role has been changing. It is a direct measurement of heparin activity in clinical samples. Furthermore, it measures the ability of heparin-bound antithrombin to inhibit a single enzyme, Factor Xa.8 If heparin is present in the specimen, it will reduce the amount of color formed.8,22 Therefore, the quantity of chromophore released is inversely proportional to the activity of heparin present.

The American College of Chest Physicians (ACCP) in its ninth edition of Antithrombotic Therapy and Prevention of Thrombosis Guidelines (2012) does not explicitly recommend a monitoring method for parenteral heparin; nevertheless, chromogenic anti-Factor Xa assay is becoming a preferred assay for monitoring UFH therapy.17,23 This utilization of Heparin Xa assay could be attributed to its advantages: (1)it is relatively simple to perform and can be automated; therefore, it results could be available in a timely fashion; (2) relative lack of factors (e.g. preanalytic, analytic, biologic conditions) that affects its outcome, resulting in increased specificity; (3) testing could be done on frozen plasma which is an advantage for batch testing; (4) direct and more accurate measurement of heparin activity; (5) it is also used to monitor LMWH – recommended monitoring assay by ACCP.4,17,20,21

Like APTT, anti-Factor Xa assay has many disadvantages at this time: (1) relatively more expensive to perform; (2) limited availability especially in smaller laboratories because they lack the equipments to perform the assay; (3) the assay still requires attention to standardization particularly with respect to calibrators; (4) interference from hyperbilirubinemia and hypertriglycenemia.17,20

Bernadette V. Enderez, SBB(ASCP)CMMT(ASCP) is a clinical laboratory scientist at Hazel Hawkins Memorial Hospital and an MS student at Rush University in clinical lab management.


1. Ng, V. L. (2009). Anticoagulation monitoring. Clinics in Laboratory Medicine, 29(2), 283-304. doi:10.1016/j.cll.2009.05.003 [doi]

2. Uprichard, J., Manning, R. A., & Laffan, M. A. (2010). Monitoring heparin anticoagulation in the acute phase response. British Journal of Haematology, 149(4), 613-619. doi:10.1111/j.1365-2141.2010.08129.x [doi]

3. Marlar, R. A., & Gausman, J. (2013). The optimum number and types of plasma samples necessary for an accurate activated partial thromboplastin time-based heparin therapeutic range. Archives of Pathology & Laboratory Medicine, 137(1), 77-82. doi:10.5858/arpa.2011-0516-OA [doi]

4. Francis, J. L., Groce, J. B.,3rd, & Heparin Consensus Group. (2004). Challenges in variation and responsiveness of unfractionated heparin. Pharmacotherapy, 24(8 Pt 2), 108S-119S.

5. Liveris, A., Bello, R. A., Friedmann, P., Duffy, M. A., Manwani, D., Killinger, J. S., . . . Weinstein, S. (2014). Anti-factor xa assay is a superior correlate of heparin dose than activated partial thromboplastin time or activated clotting time in pediatric extracorporeal membrane oxygenation*. Pediatric Critical Care Medicine : A Journal of the Society of Critical Care Medicine and the World Federation of Pediatric Intensive and Critical Care Societies, 15(2), e72-9. doi:10.1097/PCC.0000000000000028 [doi]

6. Schechter, T., Finkelstein, Y., Ali, M., Kahr, W. H., Williams, S., Chan, A. K., . . . Brandao, L. R. (2012). Unfractionated heparin dosing in young infants: Clinical outcomes in a cohort monitored with anti-factor xa levels. Journal of Thrombosis and Haemostasis : JTH, 10(3), 368-374. doi:10.1111/j.1538-7836.2012.04624.x [doi]

7. Hamilton, L. A., Abbott, G. V., & Cooper, J. B. (2013). High-risk non-ST elevation acute coronary syndrome outcomes in patients treated with unfractionated heparin monitored using anti-xa concentrations versus activated partial thromboplastin time. Hospital Pharmacy, 48(5), 389-395. doi:10.1310/hpj4805-389 [doi]

8. Bonar, R. A., Favaloro, E. J., & Marsden, K. (2012). External quality assurance for heparin monitoring. Seminars in Thrombosis and Hemostasis, 38(6), 632-639. doi:10.1055/s-0032-1321954 [doi]

9. Kitchen, S., Preston, F. E., Jennings, I., Kitchen, D. P., Woods, T. A., & Walker, I. (2009). Interlaboratory agreement in the monitoring of unfractionated heparin using the anti-factor xa-correlated activated partial thromboplastin time: A rebuttal. Journal of Thrombosis and Haemostasis : JTH, 7(12), 2157-8; author reply 2178-9. doi:10.1111/j.1538-7836.2009.03616.x [doi]

10. Gouin-Thibaut, I., Martin-Toutain, I., Peynaud-Debayle, E., Marion, S., Napol, P., Alhenc-Gelas, M., & AGEPS Hemostasis Group. (2012). Monitoring unfractionated heparin with APTT: A french collaborative study comparing sensitivity to heparin of 15 APTT reagents. Thrombosis Research, 129(5), 666-667. doi:10.1016/j.thromres.2011.11.016 [doi]

11. Vardi, M., Laor, A., & Bitterman, H. (2009). Activated partial thromboplastin time monitoring in patients receiving unfractionated heparin for venous thromboembolism in relation to clinical outcomes. Thrombosis and Haemostasis, 102(5), 879-886. doi:10.1160/TH09-06-0404 [doi]

12. Guervil, D. J., Rosenberg, A. F., Winterstein, A. G., Harris, N. S., Johns, T. E., & Zumberg, M. S. (2011). Activated partial thromboplastin time versus antifactor xa heparin assay in monitoring unfractionated heparin by continuous intravenous infusion. The Annals of Pharmacotherapy, 45(7-8), 861-868. doi:10.1345/aph.1Q161 [doi]

13. Rosborough, T. K. (1999). Monitoring unfractionated heparin therapy with antifactor xa activity results in fewer monitoring tests and dosage changes than monitoring with the activated partial thromboplastin time. Pharmacotherapy, 19(6), 760-766.

14. Vandiver, J. W., & Vondracek, T. G. (2013). A comparative trial of anti-factor xa levels versus the activated partial thromboplastin time for heparin monitoring. Hospital Practice (1995), 41(2), 16-24. doi:10.3810/hp.2013.04.1022 [doi]

15. Hartman, S. K., & Teruya, J. (2012). Practice guidelines for reversal of new and old anticoagulants. Disease-a-Month : DM, 58(8), 448-461. doi:10.1016/j.disamonth.2012.04.003 [doi]

16. Dittus, C., & Ansell, J. (2013). The evolution of oral anticoagulant therapy. Primary Care, 40(1), 109-134. doi:10.1016/j.pop.2012.11.011 [doi]

17. Wool, G. D., Lu, C. M., & Education Committee of the Academy of Clinical Laboratory Physicians and Scientists. (2013). Pathology consultation on anticoagulation monitoring: Factor X-related assays. American Journal of Clinical Pathology, 140(5), 623-634. doi:10.1309/AJCPR3JTOK7NKDBJ [doi]

18. Hirsh, J., & Raschke, R. (2004). Heparin and low-molecular-weight heparin: The seventh ACCP conference on antithrombotic and thrombolytic therapy. Chest, 126(3 Suppl), 188S-203S. doi:10.1378/chest.126.3_suppl.188S [doi]

19. Mismetti, P., & Laporte, S. (2010). New oral antithrombotics: A need for laboratory monitoring. for. Journal of Thrombosis and Haemostasis : JTH, 8(4), 621-626. doi:10.1111/j.1538-7836.2010.03764.x [doi]

20. Olson, J. D., Arkin, C. F., Brandt, J. T., Cunningham, M. T., Giles, A., Koepke, J. A., & Witte, D. L. (1998). College of american pathologists conference XXXI on laboratory monitoring of anticoagulant therapy: Laboratory monitoring of unfractionated heparin therapy. Archives of Pathology & Laboratory Medicine, 122(9), 782-798.

21. Vandiver, J. W., & Vondracek, T. G. (2012). Antifactor xa levels versus activated partial thromboplastin time for monitoring unfractionated heparin. Pharmacotherapy, 32(6), 546-558. doi:10.1002/j.1875-9114.2011.01049.x [doi]

22. Rosenberg, A. F., Zumberg, M., Taylor, L., LeClaire, A., & Harris, N. (2010). The use of anti-xa assay to monitor intravenous unfractionated heparin therapy. Journal of Pharmacy Practice, 23(3), 210-216. doi:10.1177/0897190010362172 [doi]

23. Garcia, D. A., Baglin, T. P., Weitz, J. I., Samama, M. M., & American College of Chest Physicians. (2012). Parenteral anticoagulants: Antithrombotic therapy and prevention of thrombosis, 9th ed: American college of chest physicians evidence-based clinical practice guidelines. Chest, 141(2 Suppl), e24S-43S. doi:10.1378/chest.11-2291 [doi]

About The Author