1. Anticoagulation Monitoring Shouldn’t We Validate?
Mark T. Lucas, MPS, CCP, PBMT
Chief of Perfusion
Denver Cardiovascular Perfusionists
Educational meetings should educate. They should also make you question what you do and ask if you are doing enough, the right thing or if you can do better.

2. “It’s not until you know better that you can do better.”
To solve a problem, you need to understand all of the variables.
Peter V Schiro

3. “first law of hemostasis and thrombosis.”
“The more thrombin the less bleeding but the more thrombosis”
“The less thrombin the less thrombosis but the more bleeding”
H. Coenraad Hemker, Raed Al Dieri and Suzette Béguin, Thrombin generation assays: accruing clinical relevance Curr Opin Hematol 11:170–175.

4. Current and Future Medical Practice
Trend towards evidenced based coagulation and anticoagulation management
POCT has typically been performed with less than the stringent regard for accuracy and precision than in the laboratory
Goal directed anticoagulation is the future

5. Goal Directed Anticoagulation
Focused purpose to inhibit thrombin potential
Maximizing anticoagulation at specific time points for different patient categories
Defined anticoagulant management program individualized for each patient receiving anticoagulant
Use of multiple specific anticoagulants
Unfractionated heparin
Direct thrombin inhibitors

6. Verification and Validation
..are independent procedures that are used together for checking that a product, service, or system meets requirements and specifications and that it fulfills its intended purpose.
Brian Bull ACT 480 seconds 1975 (visible clot)
John Young ACT >400 seconds 1978 (fibrin monomer)

7. Anticoagulation Effectiveness
Goal of anticoagulation is to suppress thrombin.
How effective is your anticoagulation protocol at suppressing thrombin?
100%, 90%, 80%, 70%...How do you know?
Have you validated your anticoagulation protocol in your system?

8. DCP Anticoagulation Protocol
Arterial blood sample for baseline ACT and HMS Heparin Dose Response (500 s)
Administer unfractionated heparin via central venous catheter per HDR dose
Arterial blood sample for ACT after 3–5 min
Ensure ACT 3–4 times above baseline ACT (>400 s) before initiating CPB
10,000 IU unfractionated heparin in CPB prime solution
Start continuous heparin infusion with initiation of bypass at 100 IU/kg/hr
First on-bypass sample within ten minutes of initiation, then monitor ACT/HPT 30 min during CPB <35 degrees and 20 min >35 degrees
Maintain HPT +/- one channel from target and ACT >400 s
Reverse heparin with protamine per HPT value after separation from CPB. Dose ratio .8 mg protamine per 1000 IU of heparin
Arterial blood sample for ACT/HPT 3–5 min post protamine

9. Variability in Dosing 74% empiric dosing 20% heparin dose response
5% fixed number
1% other

10. Variability in Maintenance
ACT value
ACT/Protamine Assay
Set time intervals
Continuous infusion

11. Variability in Testing Method
59% of monitoring by ACT alone
33% ACT and Protamine titration
2% Viscoelastic
6% Other

12. Variability in Testing Frequency
Testing every 10 minutes to greater than 60 minutes between tests.
Or no testing at all.

13. Variability of Minimum ACT
5% Other

14. Goals of Anticoagulation
Target ACT
Preserve Coagulation factors
Inhibit visible clot
100% Thrombin inhibition
Target protamine titration

15. Chromogenic Anti-Xa Test
The factor Xa assay is really a drug level using chromogenic Xa inhibition as the methodology; as the reference range and standard curve vary with the drug tested.
Chromophore-linked substrate of factor Xa. Upon cleavage of the substrate by the active enzyme (factor Xa), a colored compound is released

16. How Xa Assay Works
Known amounts of factor Xa and antithrombin added to the sample
Heparin forms an inhibitory complex with antithrombin and inactivates factor Xa
Excess amount of factor Xa remaining in the sample is inversely proportional to the amount of heparin
Results are then compared to a standard curve and are provided in concentration of anti–factor Xa (units/mL).

17. Thrombin Generation Testing
20 patients – 5 CABG, 5 Valve, 5 DM, 5 HR
Samples for analysis concordantly with Heparin protocol:
Pre-bypass (baseline)
Post heparin bolus (non-stimulated)
10” post bypass (stimulated)
10” post x-cl removal (stimulated)
Post protamine dose (stimulated)

18. Validation Testing Goal
Determine amount of thrombin produced for time points during procedure known for minimal and maximal thrombin stimulus
Pre-surgery
Post heparin bolus
Post initiation bypass
Post cross clamp removal
Post heparin reversal with protamine

19. Thrombin Generation Assay (Currently for research use only)
Calibrated Automated Thrombogram (CAT)
Whole blood samples
Flurigenic substrate
Tissue factor stimulus
Measures initiation of thrombin, fibrin formation, phase down and inactivation
All forms of anti-coagulant therapies such as heparin therapy, direct thrombin inhibitors or oral anti-coagulants can be monitored
TECHNOTHROMBIN® TGA is based on monitoring
the fluorescence generated by thrombin cleavage of
a fluorigenic substrate over time upon activation of
the coagulation cascade by different concentrations
of tissue factor (RA, RB, RC Low, RC High and RD*)
and negatively charged phospholipids in plasma.
From the changes in fluorescence over time, the
concentration of thrombin (nM) in the sample can be
calculated using the respective thrombin calibration
curve (only one calibration has to be done for each
lot). The rate of increase in thrombin concentration
over time then allows to calculate generation of
thrombin in the sample per minute and to plot this
value over time for the whole coagulation process.
This then results in the visualization of the different
phases of clot formation.

20. Balance
Balance between anticoagulation for cardiopulmonary bypass (CPB) and hemostasis after CPB.
Too much heparin – bleeding
Too little heparin – clot/bleeding

21. Summary Thrombin inhibition is the primary goal
Considerable variability in anticoagulation practice and performance
Technology affords us new opportunities
Validation of anticoagulation effectivenesss is necessary for consistency and improved outcomes in CV surgery globally