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Assessing Platelet Reactivity with Point-of care Systems
Christian Valina Herz-Zentrum Bad Krozingen
Oral Antiplatelet Therapy After PCI: Optimizing Outcomes in Clinical Practice
Assessing Platelet Reactivity with Point-of care Systems
Platelet-function tests
What kind of POC-tests are available?
Differences in test results
Assessing Platelet Reactivity with Point-of care Systems
Platelet-function tests
What kind of POC-tests are available?
Differences in test results
Platelet-function tests
Test principle TestsCessation of bleeding from a standardized wound in vivo
Bleeding time
Aggregometry Light transmission aggregometry (LTA)Multiplate® WBAVerifyNow®
Platelet counting method (e.g. Ichor-plateletworks®)Thrombovision T® guide
Flow cytometry Detection of activation markers ex vivoEx vivo stimulation and detection of activation markersVASP phosphorylation
Shear dependent platelet function within whole blood
PFA-100®
IMPACT® -CPAPlacor PRT-7000®
Global haemostasis tests Thrombelastography (TEG®)Thrombelastometry (ROTEM®)Haemostasis Analysis System®
Thromboxane measurements Serum thromboxane B2
Harrison, Hämostaseologie 2009; 29: 25
Platelet function testing: Some Problems
• No definitive screening test
• Methodological variability within each technique
• Labor intensive
• Costly
• Time consuming
• Requirement of fair degree of expertise and experience to perform and interpret
• Often requirement of additional expensive specialist tests
• Lack of a standardized definition of suboptimal platelet response
Assessing Platelet Reactivity with Point-of care Systems
Platelet-function tests
What kind of POC-Devices are available?
Differences in test results
Point-of-care-testing: What should they can?
• Bed-side testing
• No sample preparation
• Ready-to-use
• Easy to use
• Rapid read out
• Short Turn-around-Time (TAT)
Point-of-care-testing: Test Principles
POC-System Test principle
VerifyNow®
Multiplate® WBAAggregometry
PFA-100®
IMPACT® -CPAPlacor PRT-7000®
Shear dependent platelet function within whole blood
Thrombelastography (TEG®)Thrombelastometry (ROTEM®)Haemostasis Analysis System®
Global haemostasis tests, monitoring of rate and quality of clot formation
Ichor-Plateletworks® Platelet counting pre- and postactivation
HemoStatus® Platelet procoagulant activity
VerifyNow®
• Whole blood (citrated) without further sample preparation
• No need for time-consuming centrifugation steps
• Turbidimetric based optical detection of platelet aggregation stimulated by a specific agonist
• Disposable cartridges contain fibrinogen-coated beads and platelet activator (3 cartridges available: aspirin, P2Y12, GPIIb/IIIa)
Results Screen VerifyNow P2Y12 Assay
PRU result is ‘P2Y12-mediated platelet aggregation’ via
adenosine diphosphate (ADP) pathway
VerifyNow® P2Y12 results can then bereported as % platelet inhibition
Base result is ‘Maximal plateletaggregation’ via Thrombin ReceptorActivating Peptide (TRAP) pathwaywhich is independent of aspirin and
clopidogrel
Correlation of VerifyNow® with LTA
0 20 40 60 80 1000
100
200
300
400
500
r2= 0.434p< 0.0001n=562
RPA ADP 5 µM, %
Ve
rify
No
w P
2Y12
PR
U
Multiplate® system (“multiple platelet function analyzer”)
• Whole blood (hirudinized)• Multiple electrodes in the disposable test cell (4 electrodes form 2 independent
sensor units), multiple channels of the instrument (5).• Detects the effects of the platelet inhibitors Aspirin®, clopidogrel and GpIIb/IIIa
antagonists, and also the newer direct ADP receptor antagonists.• No luminescence available, low shear system, does not simulate normal
haemostasis.• Continuously records platelet aggregation.• Increase of impedance by the attachment of Platelets onto the Multiplate® sensors
is transformed to arbitrary aggregation units (AU) and plotted against time. • Three parameters: – Area under the aggregation curve (AUC).– Aggregation is the height of the curve. V– Velocity is the maximum slope of the curve.
Platelet Function Assay 100 (PFA-100)
• Analyzer device and disposable cartridge
• Whole blood
• Measurment of time to occlusion of aperture in mebrane coated with collagen/ADP or collagen/epinephrin under high shear stress conditions
• Cannot distinguish thienopyridene effect
160µm- 40mbar
Flow direction
Thromboelastography (TEG) Haemoscope's TEG® 5000 system
• Small sample of blood (typically 0.36 ml), is placed into a gently rotating cuvette (imitates venous flow and activate coagulation) => Sensor shaft inserted into the sample => a clot forms between the cup and the sensor.
• Speed and strength of clot formation is measured.• Patterns of changes in strength and elasticity in the clot provide information.• Four values that represent clot formation are determined by this test:
– the R value (or reaction time): represents the speed of clot formation – the K value: end or R until the clot reaches 20mm – the angle and
– the MA (maximum amplitude). clot strength • Coagulation Index (CI) (or overall
assessment of coagulability).
Assessing Platelet Reactivity with Point-of care Systems
Platelet-function tests: Advantages of POC-Systems
Platelet-function tests: What kind of POC-tests are available?
Differences in test results
Differences in test results: Are they measuring the same?
• Low shear systems (e.g. Multiplate) does not simulate normal haemostasis
• Different stimulation reagents and dosages
• Is „baseline aggregation“ really „baseline“?
• „Units“:– % platelet inhibition or P2Y12 reaction units (VerifyNow)
– Arbitrary aggregation units (Multiplate)
– Closure time (PFA-100)
– Maximum aggregation, residual aggregation, … (LTA)
– Platelet reacitivity index (VASP-P)
• Cut-off values
Assessing Platelet Reactivity with Point-of care Systems
Platelet-function tests: Advantages of POC-Systems
Platelet-function tests: What kind of POC-tests are available?
Differences in test results
Cut-off values
VerifyNow: Cut off levels for „low response“
• Trenk, 2008; TRIGGER (enrolling) > 208 PRU
• Godino, 2009 > 213 PRU
• Price et al., 2008 ≥ 235 PRU
• ARMYDA-PRO, GRAVITAS (enrolling) ≥ 240 PRU
• Paniccia et al., 2007 ≥ 264 PRU
Assessment of On-Clopidogrel Platelet Reactivity by LTA and VerifyNow
17.6 ± 16.2 %n=307
17.6 ± 16.2 %n=307
161.6 ± 90.5n=307
161.6 ± 90.5n=307
LTALTA VerifyNowVerifyNow
Ver
ifyN
ow
® P
RU
Ver
ifyN
ow
® P
RU
n=307r2=0.441n=307
r2=0.441
Assessment of On-Clopidogrel Platelet Reactivity Correlation Between LTA and VerifyNow
AUC: 0.833Sensitivity for PRU 208: 0.53Specificity for PRU 208: 0.88
AUC: 0.833Sensitivity for PRU 208: 0.53Specificity for PRU 208: 0.88
Assessment of On-Clopidogrel Platelet Reactivity LTA versus VerifyNow by ROC Analysis