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PubMed2005

Platelet function tests and flow cytometry to monitor antiplatelet therapy.

There are numerous methods to measure platelet function in specialized laboratories. They provide valuable results for research or small patient studies, but none of these tests is currently used to guide antiplatelet therapy in routine clinical practice. Aggregometry is the most commonly used test, but this technique is time-consuming and pre-analytical preparation, choice of anticoagulant, and agonist have not been standardized. The Rapid-Platelet-Function-Analyzer uses whole blood to measure glycoprotein (GP) IIb/III inhibitor activity. This test is quick and simple and results for abciximab correlate closely with aggregometry. The platelet function analyzer PFA-100 is a good screening method to detect platelet function defects. The closure time of the PFA is also sensitive for aspirin-resistance that affects many patients. Flow cytometry can monitor a whole panel of platelet activation markers and platelet interactions with other cells. It can quantitate receptor expression and inhibition. Although cytometry is time-consuming and expensive, it is a valuable tool for research. Platelets have many different functions. This requires specific tests optimized for each of the different functions. Each test has advantages and disadvantages and should be interpreted only in concert with a thorough history and physical examination. The ideal test should be simple, quick, inexpensive, and available on a 24-hour basis.

Abciximab↗
PubMed2001

In vitro dose response to different GPIIb/IIIa-antagonists: inter-laboratory comparison of various platelet function tests.

AIMS: The aim of this study was to assess the inter- and intra-laboratory variation of the concentration-response to the GPIIb/IIIa-antagonists abciximab and eptifibatide on platelet aggregometry and to compare results with flow cytometric tests as well as the rapid platelet function analyser (RPFA). METHODS: In five different laboratory sites, blood from three to five healthy donors was spiked with abciximab or eptifibatide, followed by the assessment of: (1) aggregometry (anticoagulant: sodium citrate 3.18% or hirudin 5 microg/ml); (2) flow cytometry (fibrinogen binding or PAC1-expression), or (3) RPFA. Dose-response curves were established on the basis of a sigmoidal Imax)-model [I=(Imax)*Cg)/(IC50g + Cg)]. RESULTS: For citrated blood, aggregation induced by 20 microM ADP was blocked up to 100% by both GPIIb/IIIa-antagonists, IC50 values varied between 0.11-0.22 microg/ml for eptifibatide and 1.25-2.3 microg/ml for abciximab. I(max) of the response to 5 microg/ml collagen ranged from 46% to 100%, and IC50 values varied between 0.28-0.34 microg/ml for eptifibatide and 2.3-3.8 microg/ml for abciximab. In hirudinized blood, IC50 values for eptifibatide were 1.5- to 3-fold higher than those obtained with citrated plasma. Inhibition of PAC1-expression by abciximab (IC50) 0.84 microg/ml) showed results similar those of the RPFA (approx. 1.0 microg/ml); larger differences between PAC1 and RPFA results were observed for eptifibatide. Based on aggregometry, eptifibatide concentrations for 80% inhibition varied from 0.27 to 0.55 microg/ml, and were considerably less when the RPFA was taken as basis (0.15 or 0.22 microg/ml). A similar pattern was observed for abciximab. CONCLUSIONS: We found quite a low inter- and intra-laboratory variation in the in vitro pharmacodynamic characterization of GPIIb/IIIa-antagonists by aggregometry, making results of these tests obtained from different laboratories during clinical trials at least comparable. The RPFA exhibits a higher sensitivity to inhibitory GPIIb/IIIa-effects, in keeping with the "real" inhibition of the activated receptor (PAC1) as assessed with more elaborate flow cytometry.

Abciximab↗
PubMed2004

The predictive value of megakaryocytic and erythroid colony formation and platelet function tests on the risk of thromboembolic and bleeding complications in essential thrombocythaemia.

The predictive value of spontaneous in vitro colony formation of megakaryocytic and erythroid progenitors (154 patients), and defective platelet aggregation responses (55 patients) on the risk of thrombohaemorrhagic complications in patients with essential thrombocythaemia (ET) was evaluated retrospectively. In the in vitro cultures of haematopoietic progenitors, 114/154 patients (74%) showed either spontaneous megakaryocytic or erythroid colony formation or both. Forty-three per cent of patients with any spontaneous colony growth and only 20% of those without this phenomenon had an arterial thrombosis at diagnosis or during the follow-up (P = 0.02). In the whole patient group neither spontaneous megakaryocytic nor spontaneous erythroid colony formation alone predicted the risk of arterial thrombosis. In patients younger than 45 yr of age, the prognostic value of spontaneous megakaryocytic growth was statistically significant: 44% of the patients with spontaneous megakaryocytic colony formation, but only 14% of those without it, experienced arterial thrombosis (P = 0.04). The presence of spontaneous colony formation had no effect on the risk of bleeding complications. Forty-one of the 55 patients (75%) showed abnormalities in the platelet aggregation responses. There was no statistically significant correlation between the platelet function response and the risk of bleeding or thrombotic complications. No correlation was found between the platelet aggregation responses and the presence of spontaneous colony growth. In conclusion, spontaneous colony formation indicated an increased risk of thrombohaemorragic events but the platelet function test had no predictive value for these complications.

Adult↗
PubMed1982

The effect of prednisone on platelet function tests.

Ivy bleeding time, capillary fragility, threshold ADP concentration for secondary platelet aggregation and platelet adhesiveness were found to be unchanged by 2 d and 6 weeks of treatment with prednisone in 22 consecutive patients with collagen or haematological diseases. Initial high platelet counts were unchanged after 2 d of treatment, but after 6 weeks of treatment the increase was significant. It is concluded that in patients of this category a 6-week treatment with commonly used doses of prednisone does not significantly affect platelet function.

Antigens↗
PubMed1989

A sensitive in vivo platelet function test in rats based on intravenously injected collagenase.

The transient decrease of the platelet concentration in the flowing blood after intravenous injection of collagenase was used as a measure for the effectiveness of the platelet-vessel wall interactions in rats. The decrease of the concentration in circulating platelets was regarded as an expression of the in vivo platelet function. The influence of acetylsalicylic acid, imidazole, indomethacin, ketanserin, RA 233, and dipyridamole on this test was investigated in anaesthetized rats. All tested drugs showed an effect on the extent of the platelet concentration decrease after the collagenase injection compared to the control values, but no drug was significantly effective. This test could prove to be a valuable tool for testing the efficiency of antithrombotic drugs in vivo.

Animals↗
PubMedDate not supplied

Platelet function testing by flow cytometry.

The availability of fluorescent monoclonal antibodies and probes now provides a powerful tool for examining platelet function by flow cytometry. These techniques cna be employed to determine the resting and stimulated expression of platelet glycoprotein receptors, the activation status of platelets assessed by secretion of granule contents (including expression of activation-dependent neoantigens), adhesion ligand binding to platelets, and intracellular calcium flux after exposure of platelets to agonist. In addition, flow cytometry has now been used to study the functional properties of stimulated platelets, including microparticle generation, platelet aggregation, and platelet-heterotypic cell conjugate formation. This brief review is presented as a general outline of the literature that uses flow cytometric methodology to examine in vivo and ex vivo platelet function.

Blood Platelet Disorders↗
PubMed1989

Whole blood aggregometry. Influence of sample collection and delay in study performance on test results.

The method of sample collection and time interval from venipuncture to test performance is considered to be important in platelet function testing. Platelet function studies can be performed in whole blood with the use of an impedance lumi-aggregometer, and an examination of the influence of these variables on the outcome of such a study is important. Twelve healthy donors had blood drawn with the use of conventional approach for platelet function testing and a Vacutainer method. Platelet function studies were performed at three time points over a three-hour period. No significant differences in results were observed between the methods of collection and the time to test performance. It is concluded that for platelet function screening in whole blood, with the use of an impedance lumi-aggregometer, a Vacutainer sample maintained at room temperature and tested within three hours is satisfactory.

Adenosine Triphosphate↗
PubMed2004

Plateletworks platelet function test compared to the thromboelastograph for prediction of postoperative outcomes.

Approximately 3.5 million units of platelets are transfused in the United States each year to patients undergoing open-heart surgery with cardiopulmonary bypass (CPB). CPB is a known contributor to platelet loss and platelet dysfunction leading to disruption of hemostasis. Impaired hemostasis results in excess bleeding in 5-25% of all patients undergoing CPB. For this reason, it may be beneficial to measure platelet number and function in these patients. The purpose of this study was to compare the Plateletworks platelet function analyzer to the thromboelastograph (TEG) in predicting postoperatiave hemostatic outcomes as measured by blood product use and chest tube (CT) drainage. This study consisted of 35 adult patients undergoing cardiac surgery with cardiopulmonary bypass at Rush-Presbyterian-Saint Luke's Medical Center (RPSLMC). The Plateletworks and TEG tests were performed preoperatively, after protamine was given, and 24 hours postoperatively on all patients. Plateletworks demonstrated a statistically significant change in platelet function as shown by the adenosine diphosphate (ADP) reagent tube from the preoperative period to the removal of the aortic cross clamp (p = .011). The TEG did not demonstrate a significant change in the k-time and maximum amplitude (MA), but did show a significant change in the alpha-angle from the pre-operative to postoperatiave sample (p = .035). A correlation was found between Plateletworks collagen reagent tubes preoperatively and CT drainage (p = .048, r -0.324). No statistical correlation was established between TEG parameters and CT drainage at any time interval. TEG preoperative MA showed a correlation to receipt of blood products (p = .016). When comparing the Plateletworks to the TEG in this study, the Plateletworks system was a more useful predictor of blood product use and chest tube drainage.

Adult↗