Emergency and Perioperative Hemostasis Testing: Which Assays Provide Helpful Information. Wayne Chandler, MD Laboratory Medicine Seattle Children s

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1 Emergency and Perioperative Hemostasis Testing: Which Assays Provide Helpful Information Wayne Chandler, MD Laboratory Medicine Seattle Children s

2 Emergency Hemostasis Testing Patients actively bleeding Trauma: the Golden Hour Intra and Post-operative bleeding Obstetrical bleeding GI hemorrhage Ruptured aortic aneurysm Intracranial hemorrhage

3 Hemostasis testing in the bleeding patient What is wanted: Rapid assessment to determine which blood products are needed: FFP, Factor Concentrates, Platelets, Cryoprecipitate What isn t wanted Super accurate measurement of hemostatic parameters that takes an hour to complete

4 Hemostasis testing in the bleeding patient Speed is critical, slow testing may be of no clinical benefit if it arrives too late Accuracy is important, but focus should be on accuracy sufficient to make clinical decisions within the shortest time

5 Emergency Testing, Clinical Lab First Step, Meetings with Users Meet with surgeons, anesthesiologists, others in need of emergency hemostasis testing Rules Must limit testing to actively bleeding patients Must strive to get a good sample Their responsibility to deliver sample Must select method for result delivery e.g. provide phone # Lab focus on speed, sufficient accuracy 1:1 correspondence between result and treatment Red light Green light factors low, transfuse factors adequate, don t transfuse

6 Emergency Testing, Clinical Lab Emergency Assessment Test Methodology Options Standard Hematology/Coagulation Test PT, Fibrinogen, Platelet Count, Hematocrit Viscoelastic Whole Blood Assays TEG, Rotem Emergency Platelet Function Testing PFA, VerifyNow

7 Emergency Testing in the Clinical Laboratory Typical Diagnostic Coagulation Screen Focus is on screening for possible acquired or hereditary hemostatic defects that may explain past problems and predict future risk of bleeding. Assumption is the patient is not actively bleeding at present. Time is important, but accurate final diagnosis is more important: vwd, platelet disorder, lupus inhibitor, elevated or reduced factor VIII, etc Hematology Clinic

8 Standard Stat Coag Screen PT, PTT, Thrombin Time, Fibrinogen, Platelet Count, D-dimer Requires minutes Rejected samples Focus on wide range of possible coag abnormalities, congenital deficiency, acquired abnormalities, inhibitors, etc. Chandler et al. Transfusion 2010;50:2547

9 Sample Processing Expedite Central Processing Announce Emergency Hemorrhage Panel Check sample against order, if they match Move sample immediately to Heme/Coag Can be logged in parallel with testing Use rapid 2 min spin centrifuge

10 Factors evaluated by PT and PTT The PT provides a better estimate of coagulation factor levels in the bleeding patient. It is faster than PTT It suffers fewer problems than PTT with interference from high factor VIII, contact system issues and lupus inhibitors Good correlation with average factor level and response to FFP PT 84% sensitivity (detect factor <35%) PTT 50% sensitivity Yuan, Chandler, Thr Res 2007;120:29

11 Rapid fibrinogen assay Coagulation analyzer programmed to measured fibrinogen between ~150 and 900 mg/dl. Fibrinogen below 150 mg/dl caused repeat assays and dilutions which delayed result up to 24 minutes. Reprogrammed instrument to use a modified calibration curve to result all fibrinogens between 50 and 1100 mg/dl

12 Effect of hemolysis on PT and Fibrinogen 3% of samples hemolyzed. Compared PT and fibrinogen on 67 hemolyzed samples vs replacement samples Hemolyzed Unhemolyzed P PT 23.3± ±10.2 NS Fib 458± ± Difference not considered clinically significant

13 Effect of clotting on PT and Fibrinogen 1% of samples clotted. Compared PT and fibrinogen on 24 samples with a clot or fibrin strands detected versus replacement samples Clotting No clot P PT 14.9± ± Fib 366± ± PT change not considered clinically significant. Only one clotted sample had a fibrinogen less than 100 mg/dl, error of about 1 in 2300 samples.

14 Focus on: Speed Factor deficiency due to bleeding Each assay directed at specific blood product Chandler et al. Transfusion 2010;50:2547

15 Accuracy vs Time In vitro clotting and platelet clumping can lead to falsely low fibrinogen and platelet counts, but If the true fibrinogen was 400 mg/dl and the sample fibrinogen 300 mg/dl would you treat the patient differently? Is it better to let the patient care team know the fibrinogen is at least 300 mg/dl now, versus ask for a new sample? Think of the results as: The fibrinogen is at least *** The platelet count is at least *** This is often all the physician needs for making transfusion decisions.

16 One approach Emergency Hemorrhage Panel For speed, run the sample once PT, fibrinogen, platelet count, hematocrit If fibrinogen <100 mg/dl or platelet count <100,000/uL check for clot and let team know if result may be falsely low Immediately report result to ordering location We detected false low in about 1:2000 samples Available in about min Chandler et al. Transfusion 2010;50:2547

17 Whole blood viscoelastic testing for hemostasis Thromboelastograph and Rotem Measures clot viscoelasticity (think clot strength) over time. Multiple parameters measured: Time to start clotting ~ PT or PTT Maximum clot strength ~ platelet count and fibrinogen Loss of clot strength ~ fibrinolysis

18 TEG and ROTEM analysis

19 Main variables TEG and ROTEM TEG R time to clot onset K time to 20 mm amplitude Angle rate of clot formation MA maximum amplitude LY30 EPL at 30 min LY60 EPL at 60 min EPL estimated % of lysis ROTEM CT - time to clot onset CFT time to 20 mm CF Angle rate of clot formation MCF maximum clot firmness LI 30 - Lysis Index after 30 min LI 60 - Lysis Index after 60 min ML - Maximum Lysis % MCF

20 Angle and Maximum Amplitude/Firmness Angle = rate of clot formation/strength MA and G = maximum clot strength Often reported Angle ~ fibrinogen Amplitude/Firmness ~ platelet count Best correlation was with platelet count and fibrinogen TEG/Rotem alone cannot distinguish fibrinogen deficiency from decreased platelets Larsen et al. Anesthesiology 2011;115:294

21 MA (Elastic Shear Modulus) vs Platelet Count r 2 = 0.70

22 Effect of hematocrit on whole blood fibrinogen assay Plasma Fibrinogen = 200 mg/dl Whole Blood same sample Fibrinogen = 100 mg/dl Amukele TK, Ferrell C, Chandler WL. Am J Clin Pathol Apr;133(4):550-6.

23 Fibrinolysis: Trauma, Cardiopulmonary Bypass and Liver Transplantation Systemic fibrinolysis is associated with an increased risk of bleeding Fibrinolysis is not easily detected with standard coagulation assays PT, PTT, Fib, etc Anti-fibrinolytic therapy is effective at reducing fibrinolysis associated bleeding Viscoelastic testing can detect severe fibrinolysis, but is slow, generally takes about an hour. Cannot detect mild/moderate fibrinolysis (PAI-1 deficiency)

24 Case Bleeding during surgery 62 year old man Undergoing cadaveric renal transplantation secondary to diabetes associated renal failure Surgery proceeding normally when surgeons noted increasing blood loss. Anesthesiologist reports new onset bleeding from all intravenous lines Blood loss accelerated and emergent RBC transfusions started Normal PT, PTT, Fibrinogen, Platelet Count

25 Viscoelastic Testing & Fibrinolysis Baseline tpa = 20 ng/ml Surgery tpa = 125 ng/ml

26 Viscoelastic Testing Emergency evaluation trauma, post-op Normal result useful, good NPV Detects severe fibrinolysis when other tests do not Detects combined low plt, low fibrinogen or low plt and fibrinogen, but not specific (low angle, MA) Relatively fast Weaker at factor and fibrinogen estimation (r 2 = 0.6) Decreases blood loss and transfusion but not morbidity or mortality

27 Emergency Evaluation of Platelet Function Pre-operative clearance for urgent surgery Unknown platelet function defects/vwd Current level or clearance of anti-platelet therapy Left ventricular assist devices/artificial hearts Anti-platelet therapy monitoring Intracranial hemorrhage Detection of anti-platelet therapy

28 VerifyNow Optical detection instrument Platelet function in the presence of Aspirin (Arachidonic acid) Clopidogrel (ADP) Am J Cardiol 2006;98[suppl]:4N 10N

29 Aspirin Testing - Thrombosis vs Bleeding VerifyNow Aspirin If ARU greater than 550 increased risk of thrombosis. More aspirin does not seem to increase inhibition If ARU less than 480 increased risk of bleeding While some platelet function tests show some prognostic value related to aspirin therapy, too much variability, no real consensus Dretzke et al. Health Technol Assess 2015;19:1-366

30 Platelet Function & TEG In a standard assay, platelets activated by thrombin. Standard TEG/Rotem are not sensitive to aspirin or clopidogrel/prasugrel TEG Platelet Mapping platelet function test that requires 4 TEG assays Problems with assay design Assay Sample Activator Plt Act Fib Act MA thrombin Citrate WB Kaolin Thrombin Thrombin MA fibrin Heparin WB Reptilase, F13 MA ADP Heparin WB ADP, Reptilase, F13 MA AA Heparin WB Arach Acid, Reptilase F13 None ADP Arachidonic Acid Reptilase, F13 Reptilase, F13 Reptilase F13

31 Platelet Mapping ADP Inhibition Plavix?? TEG Platelet Mappin Collyer T C et al. Br. J. Anaesth. 2009;102:

32 Platelet Mapping Issues Assessment of platelet inhibition secondary to clopidogrel and aspirin therapy in preoperative acute surgical patients measured by Thrombelastography Platelet Mapping T. C. Collyer et al., Brit J Anaesth 2009;102:492 CONCLUSIONS: TEG-PM can identify statistically significant platelet inhibition after antiplatelet therapy; however, the overlap in platelet receptor inhibition between the three groups is likely to limit the clinical usefulness of this test.

33 Platelet Function Testing VerifyNow There was a correlation between preoperative platelet inhibition measured by VerifyNowP2Y(12) and surgical blood loss or transfusion requirements. However, for the individual patient, preoperative use of VerifyNowP2Y(12) as an instrument to decide bleeding and transfusion risk does not seem helpful. TEG Platelet Mapping No correlation between preoperative results and blood loss or transfusion requirements Alström U et al. Thromb Res 2009;124:572

34 Platelet Mapping Issues Normal Range for inhibition Up to 70% inhibition for Arachidonic Acid (average 20%) Up to 90% inhibition for ADP (average 48%) Thrombin and Platelet Breakthrough Unexpected platelet or coag activation Results in erroneous estimates of % inhibition May require yet another special tube (5 th assay) Low Reproducibility TEG CV 67%, VerifyNow CV 5%