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Background: Dynamic adhesion assays allow the examination of platelet dysfunction and drug effects on platelet function., Objective: The purpose of the study was to optimize several parameters such as type and concentration of collagen, wall shear stress, and the concentration of the platelet-activating agonist in a new biochip perfusion chamber for the study of canine platelets., Methods: After fluorescent staining of platelets, citrated blood of 10 healthy dogs was perfused through the flow chamber coated with different concentrations of canine or bovine skin collagen. Wall shear stress ranged from 14 to 60 dynes/cm(2). Protease-activating receptor 4 (PAR 4) agonist was used for platelet activation. After perfusion, platelet attachment to the collagen matrix was quantified based on fluorescent imaging. Total platelet covered area and average size of platelet covered areas were measured by planimetry., Results: Canine platelet adhesion was supported by ≥ 200 μg/mL canine collagen, but not bovine skin collagen. Consistent results were obtained with a wall shear stress of 14 dynes/cm(2), whereas higher wall shear stress resulted in increased variability. Platelet activation with PAR 4 agonist increased the total platelet covered area and the average size of platelet covered areas., Conclusions: This study indicates the need to carefully select collagen type and concentration to assess canine thrombus formation in a dynamic flow chamber. The established method should be a useful tool to determine changes in platelet-matrix interactions as an indicator of platelet activation or platelet dysfunction in dogs., (© 2013 American Society for Veterinary Clinical Pathology.)

Objective: To determine whether soybean oil emulsion has an in vitro effect on platelet aggregation and thromboelastography in blood samples obtained from healthy dogs., Animals: 12 healthy adult dogs., Procedures: Blood samples were collected from each dog into tubes containing EDTA, hirudin, or sodium citrate for a CBC, collagen- and ADP-induced impedance aggregometry, or thromboelastography, respectively. Whole blood platelet aggregation, determined with ADP or collagen agonists, was measured in blood samples containing hirudin and final lipid concentrations of 0, 1, 10, and 30 mg/mL. The thromboelastographic variables R (reaction time), K (clotting time), α angle, and maximum amplitude were evaluated in blood samples containing sodium citrate and final lipid concentrations equivalent to those used for assessment of platelet aggregation., Results: Median maximum ADP- and collagen-induced platelet aggregation in blood samples containing 1, 10, or 30 mg of lipid/mL did not differ significantly from the value for the respective lipid-free blood sample. Maximum amplitude determined via thromboelastography was significantly reduced in blood samples containing 10 and 30 mg of lipid/mL, compared with findings for lipid-free blood samples. Values of other thromboelastographic variables did not differ, regardless of lipid concentrations., Conclusions and Clinical Relevance: Maximum amplitude determined via thromboelastography in canine blood samples was significantly affected by the addition of lipid to final concentrations that are several orders of magnitude higher than clinically relevant lipid concentrations in dogs. Lipid treatment appears to have no significant effect on hemostatic variables in dogs, although clinical studies should be performed to confirm these in vitro findings.

Objective: To evaluate the effect of 6% hydroxyethyl starch (HES) solution, with a molecular weight of 130 kDa and a degree of substitution of 0.42, on canine platelet function in vitro., Samples: Blood samples from 31 healthy adult dogs., Procedures: Citrated blood was diluted with saline (0.9% NaCl) solution or HES 130/0.42 in ratios of 1:9 (ie, 1 part saline solution or HES 130/0.42 and 9 parts blood) and 1:3. Platelet plug formation time (closure time [Ct]) was measured with a platelet function analyzer and cartridges coated with collagen and ADP., Results: Median baseline Ct with citrated blood was 84.0 seconds (interquartile range, 74.5 to 99.5 seconds). Results obtained with 1:9 dilutions with saline solution and HES 130/0.42 were not significantly different from baseline results. The 1:3 dilutions with saline solution and HES 130/0.42 resulted in median Cts of 96.0 seconds (interquartile range, 85.5 to 110.8 seconds) and 112.0 seconds (92.0 to 126.0 seconds), respectively. Results obtained with both 1:3 dilutions were significantly different from baseline results. The Ct obtained with the HES dilution was also significantly different from that of the 1:3 dilution with saline solution., Conclusions and Clinical Relevance: Saline solution and HES 130/0.42 in a 1:3 dilution affected canine platelet function by prolonging Cts. The HES 130/0.42 had a significantly greater effect on canine platelets than did saline solution.

Objective: To investigate the performance of the Multiplate platelet function analyzer with regards to: (1) the use of 3 different anticoagulants (ie, citrate, hirudin, and heparin) and (2) the evaluation of optimal assay time., Design: Prospective observational in vitro study., Setting: University veterinary teaching hospital., Animals: Twenty clinically healthy dogs and 3 ill dogs., Interventions: None., Measurements and Main Results: A total of 184 analyses were performed with duplicate measurements in each test cell and results are reported as mean of the 2 measurements. Analyses were performed on blood samples from 20 dogs collected in citrate, hirudin, or heparin. A total of 4 analyses were performed on every blood sample using adenosine diphosphate, collagen, and arachidonic acid as agonists as well as a control with 0.9% sodium chloride (buffer). Aggregation in hirudin samples was significantly increased compared with heparin at all analysis times except at 6 minutes when using ADP as agonist; however, hirudin samples also demonstrated significant aggregation in the buffer control, compared to both citrate and heparin. Citrated samples yielded significantly lower aggregation compared with both hirudin- and heparin-stabilized samples at 6 and 12 minutes when ADP and collagen were used as agonists, and at most analysis times with arachidonic acid. The assay performed best at shorter analyses times, whereas longer analyses times yielded larger variation in data., Conclusions: There was a good aggregation response and acceptable analytical variation in both heparin- and hirudin-anticoagulated samples with all tested agonist at the concentrations recommended by the manufacturer. The results suggest that heparin may be superior as anticoagulant for Multiplate analyses in dogs and that short analyses times are preferable. Spontaneous platelet autoaggregation in hirudin samples warrants careful evaluation of results using this anticoagulant, especially at longer test times. The use of citrate is discouraged for Multiplate analyses in dogs due to a weak aggregation response., (© Veterinary Emergency and Critical Care Society 2012.)

Objective: To review the current in vivo and in vitro tests of platelet function (PF) currently available and applicable to companion animals., Data Sources: Scientific reviews, case reports, original clinical and laboratory research publications, and recent veterinary research conference proceedings., Human Data Synthesis: Disorders of primary hemostasis are very common in human medicine. These include inborn errors of PF and granule storage contents, primary disease mechanisms that alter PF, disorders secondary to surgical interventions, and the effects of anticoagulant medications. Knowledge of PF disorders and the optimal method for assessment must be known to understand the mechanism and to monitor the process or drug therapy., Veterinary Data Synthesis: Interest in the study and treatment of primary coagulopathies in clinical veterinary patients has resulted in a surge of recent publications and scientific research presentations. A translational approach that uses laboratory and point-of-care tools to uncover the pathophysiologic mechanisms in the patient with defects in primary hemostasis allows the clinician to plan the diagnosis and treatment more effectively., Summary: Primary hemostatic disorders are being more commonly recognized in clinical veterinary practice. The diagnosis of platelet dysfunction may be obtained via point-of-care analyzers that use relatively small blood samples and have a quick turnaround time. Recent investigations may lead to a better understanding of the pathophysiology of PF disorders and potentially the optimization, or discovery, of novel treatments., Conclusions: The assessment of PF can be completed through in vivo and in vitro point-of-care techniques as well as by submission of blood samples to more specialized platelet biology laboratories. The information obtained including the physical examination and clinical manifestations of a hemostatic disorder, as well as the benefits of each testing modality, must be known prior to the diagnostic investigation of a patient with a coagulopathy., (© Veterinary Emergency and Critical Care Society 2012.)

Tests that evaluate many aspects of platelet function have been applied in both human and veterinary medicine for the monitoring of treatment with platelet function inhibitors and for detection of platelet function abnormalities (inherited or acquired). Interspecies variation in the response to various platelet agonists is an important consideration when methods that have been developed for people are applied in other species. At the present time, many of these assays are not readily available in standard veterinary practice. Advanced platelet function testing for veterinary patients is offered at select academic institutions. Discussion with a specialist is recommended when considering the use of these tests, and the relative strengths and limitations of each assay should be considered in the interpretation of test results.

The objective of this study was to detect the influence of short-term storage on the haemostatic function in whole citrated ovine blood at different storage temperatures. Ovine blood was collected in a commercial transfer bag system containing CPDA-1 and stored on a wobbler at room (20-25 °C; n=5) or refrigerator temperature (4 °C; n=5). The following analyses were performed initially and after 1, 2, 3, 4, 5, 6, 8, 12, 24, 48 and 72 h of storage: platelet count and (spontaneous) aggregates, agonist-induced platelet aggregation with two methods (impedance aggregometry, turbidimetric method), prothrombin time, activated partial thromboplastin time, thrombin time, fibrinogen concentration and resonance thrombography. Platelet count remained stable at room temperature, whereas a significant decrease was detected after 48 h storage at 4 °C. The latter was associated with the formation of a high percentage of platelet aggregates (50-60%) after 5h storage. Decrease in platelet aggregation was significantly more pronounced when blood was stored at 4°C. The plasmatic coagulation tests were stable within the observation period. Results indicate that platelet count and aggregability of CPDA-1-stabilised ovine blood is better preserved at room temperature and provides adequate haemostatic function for ex vivo experiments for one working day. Functional loss and high percentage of platelets within aggregates which were observed in ovine blood stored at refrigerator temperature have to be considered in blood transfusion in sheep., (Copyright © 2010 Elsevier Ltd. All rights reserved.)

The selective inhibition of platelet function in whole blood coagulation testing may allow insights into the nature of hypercoagulability in dogs with critical illness. To determine the effects of cytochalasin D and abciximab on hemostatic parameters in canine citrated whole blood, an in-vitro study was designed using thromboelastography (TEG) and a platelet function analyzer (PFA-100®). 8 clinically healthy mixed breed dogs donated blood that was anticoagulated with 3.2% sodium citrate in a 9:1 blood-to-citrate ratio. Addition of cytochalasin D to citrated whole blood from 6 dogs at concentrations ranging from 0 µg/ml to 10 µg/ml caused a maximal reduction of TEG maximum amplitude (MA) at a concentration of 7.5 µg/ml (52.7 ± 4.3 to 14.3 ± 7.8 mm). Addition of abciximab to canine citrated whole blood at concentrations of either 20 µg/ml or 40 µg/ml did not affect the TEG tracing; however, addition of abciximab to citrated canine whole blood at concentrations of 10 µg/ml and 20 µg/ml significantly prolonged PFA-100 closure times (72.5 ± 15 to 149.2 ± 91 sec and 275.6 ± 54 sec, respectively, P < 0.04). Inhibition of canine platelet function by cytochalasin D is demonstrated by TEG, but abciximab did not change TEG tracings. Abciximab does, however, inhibit platelet aggregation under shear stress as measured by the PFA-100. Inhibition of canine platelet function with cytochalasin D may allow further TEG studies in dogs with clinical disease.

The aim of this study was to examine the influence of congenital portosystemic shunts (CPSS) on primary haemostasis in dogs. Bleeding time, automated platelet function analysis (PFA 100 analyser), platelet count and platelet aggregation using different methods and agonists were measured in 10 dogs with untreated CPSS and in 10 healthy, age-matched controls. Bleeding time, platelet function analysis and platelet counts did not differ significantly between groups (P>0.05). Aggregation measured using the impedance method (area under the curve) was slightly to moderately reduced with high concentrations of collagen (e.g., 5μg/mL: 2948 ± 524 vs. 3472 ± 571 AU*Min) and arachidonic acid (e.g., 1mmol/L: 1006 ± 522 vs. 1963 ± 738 AU*Min) (P<0.05), but not with adenosine diphosphate. In contrast, collagen-induced turbidimetric aggregation revealed slightly higher maximum aggregation values in dogs with CPSS. Despite the moderately altered platelet aggregation, the lack of change in global primary haemostasis screening tests indicates that dogs with CPSS do not have regularly occurring clinically relevant disorders of primary haemostasis., (Copyright © 2010 Elsevier Ltd. All rights reserved.)

The aim of this study was to optimise the technique and establish reference values for whole blood aggregometry in dogs using a novel multiplate analyser. Measurements were performed on the hirudin-anticoagulated blood of healthy dogs using a wide range of agonists. Optimal agonist concentrations were 10 micromol/L of adenosine diphosphate, 5 microg/mL of collagen and 1 mmol/L of arachidonic acid. Ristocetin (at 0.2 and 1 mg/mL) and thrombin receptor activating peptide (TRAP-6 at 32 and 160 micromol/L) did not consistently induce platelet aggregation. Coefficients of variance for within-run imprecision (n=10 repetitions) varied from 5% to 18%. Measurement signals were significantly higher when analyses were performed on standard samples (hirudin-anticoagulated blood) compared to citrated blood or blood samples anticoagulated with citrate buffer, regardless of whether or not re-calcification was performed (P<0.05). The findings indicate that the analyser is suitable for the investigation of platelet aggregation in dogs and analysis should be performed on hirudin-anticoagulated blood using optimised agonist concentrations., (Copyright (c) 2009 Elsevier Ltd. All rights reserved.)

Background: The impact of hemolysis on thromboelastography (TEG) and platelet activation indices has not been evaluated., Objective: The aim of this study was to investigate the influence of hemolysis induced mechanically (HM) and hemolysis induced by freezing (HF) on TEG, platelet counts (PLT), and platelet activation indicators., Methods: Blood from 17 dogs was divided into the following samples: controls, HM, and HF. HM was induced by 20 repetitions of expulsion of blood through a 23 g needle. Freezing was at -80 degrees C, followed by warming to 37 degrees and dilution with equal parts room temperature blood at 22 degrees C. TEG variables that were examined included reaction time (R), coagulation time (K), angle (alpha), maximum amplitude (MA), and clot rigidity (G). Platelet indices were measured with the ADVIA 2120 hematology analyzer., Results: Hematocrit (HCT) (mean+/-SD) for controls, HM, and HF were 0.41+/-0.02, 0.39+/-0.03, and 0.25+/-0.02 L/L, respectively, consistent with decreases in HCT of 4.8% (HM) and 39.0% (HF). HM resulted in decreased R (2.5+/-0.9 minutes compared with 5.2+/-1.9 minutes for controls; P<0.001), and HF resulted in increased K (15.2+/-8.6 minutes compared with 5.3+/-4.0 minutes in controls; P<0.01) and decreased alpha (20+/-11 degrees compared with 46+/-17 degrees in controls; P<0.001). MA was decreased more in HF samples (26+/-2 mm) than in HM (38+/-8 mm) or control samples (49+/-9 mm; P<0.0001). The same applied to G values. PLT decreased after HM but not after HF. Hemolysis of both types resulted in decreased mean platelet component (MPC) concentration: control, 19.3+/-2.0, HM 15.5+/-3.4, and HF 14.3+/-0.7 g/dL (P<0.0001)., Conclusion: In hemolyzed samples decreased MPC and R suggested activated primary and secondary hemostasis, respectively, but decreased MA and G indicated reduced clot firmness, possibly due to hyporeactive platelets. TEG and platelet activation indices should be interpreted cautiously after hemolysis.

Background: Whole blood platelet aggregometry (impedance) is an important method to investigate platelet function disorders. Examination of hemostatic function in sheep is important with respect to their role as an animal model of human disease., Objective: The aim of this study was to evaluate and optimize selected methodological aspects (anticoagulant, agonist concentration) of impedance aggregometry in ovine blood using the new Multiplate 5.0 analyzer., Methods: Blood samples were collected in hirudin anticoagulant from 40 clinically healthy sheep. Samples from selected sheep were collected in citrate, with or without the addition of calcium chloride. The agonists adenosine diphosphate (ADP), collagen, ristocetin, arachidonic acid, and thrombin receptor-activating peptide (TRAP) were added in several concentrations to induce aggregation., Results: Based on maximum aggregation values and internal precision, no significant difference was found between ADP concentrations of 3-10 micromol/L and collagen concentrations of 3-5 microg/mL (P>.05). The lowest interindividual variation of approximately 3-4-fold was seen with 4 and 5 micromol/L ADP and 4 and 5 microg/mL collagen. Ristocetin, arachidonic acid, and TRAP did not induce significant aggregation at any concentration. Aggregation results were significantly lower when measured in citrate- vs hirudin-anticoagulated blood, regardless of the presence of calcium chloride., Conclusions: Our results indicate that the multiplate impedance aggregometer is suitable for the measurement of platelet aggregation in sheep using optimal agonist concentrations of 4-5 micromol/L ADP and 4-5 microg/mL collagen. Hirudin-anticoagulated blood is the preferred sample material.

The purpose of this study was to examine the influence of cyclic combination chemotherapy on primary haemostasis in dogs with malignant lymphoma. Seventeen dogs receiving cytostatic treatment for high-grade lymphoma were included in the study. The dogs were treated with a Madison-Wisconsin derived protocol, which included asparaginase, vincristine, doxorubicin and prednisolone. At different time points during the first 4 weeks of induction, platelet count, capillary bleeding time, analysis of the platelet function using the platelet function analyser PFA-100, and platelet aggregation by the Born-method were measured. The most obvious changes were found for median values of the platelet count, which increased significantly from 210,000/microL before induction to 349,000/microL during the second week of induction (P=0.0010). Median platelet count subsequently decreased by the fourth week of treatment (Friedman-test: P<0.0001). None of the parameters of platelet function (capillary bleeding time, automatic platelet function analysis, aggregation maximum) showed significant changes with time (P>0.05, Friedman-test). The results did not suggest that significant platelet dysfunction was induced by the chemotherapeutic protocol used in the study., (2009 Elsevier Ltd. All rights reserved.)

Background: The platelet function analyzer (PFA)-100 is a point-of-care instrument previously evaluated in humans and dogs. In both species, artificially prolonged platelet closure time (CT) occurs with anemia. Reliability of the analyzer in dogs becomes a concern when the HCT is between 0.25 and 0.35 L/L., Objective: The objective of this study was to further define the level of HCT at which CT is prolonged, using in vitro diluted canine blood., Methods: Citrated whole blood samples were collected from 22 healthy dogs. Initial HCT was determined and autologous platelet-rich plasma was added to samples to achieve HCTs of 0.33, 0.30, and 0.27 L/L. CT was determined in duplicate on the PFA-100 using collagen/adenosine-5'-diphosphate cartridges., Results: Compared with the initial CT in samples with HCT 0.39-0.54 L/L (CT mean+/-SD=57.8+/-5.75 seconds), significantly prolonged CTs were found in hemodiluted samples with HCT 0.33 L/L (61.1+/-4.64 seconds), 0.30 L/L (64.3+/-6.79 seconds), and 0.27 L/L (70.8+/-7.90 seconds) (P=0.029; repeated measures ANOVA)., Conclusion: Although statistical differences were found, further studies are needed to determine the clinical significance of the mild prolongation in CT associated with mild anemia. Until then, dogs with HCTs slightly <0.35 L/L should be evaluated cautiously for platelet dysfunction using the PFA-100.

The objective of the study was to investigate the value of additional tests [platelet count, partial thromboplastin time (PTT), platelet function analysis using the PFA-100, Collagen binding assay (vWF:CBA), and Factor VIII activity], for use in conjunction with the von Willebrand factor antigen enzyme-linked immunosorbent assay (ELISA), as part of a newly developed diagnostic profile for improved characterization of patients with von Willebrand disease (vWD). The study population included 183 clinically healthy canines ranging in vWF:Ag concentration from 1% to 125%. The Asserachrom vWF:Ag ELISA assay was used as an external control for the determination of vWD status. Degree of association between the additional tests and vWF concentration was evaluated, and associations between the additional tests were also assessed, including their ability to distinguish dogs with vWD from those without vWD. In addition, a reference interval was determined for the PFA-100 platelet function analyzer. Strong associations were found between the PFA-100, vWF:CBA, and Asserachrom vWF:Ag assay, and a significant association was found between the PFA-100 and vWF:CBA. An association was detected between Factor VIII activity and the Asserachrom vWF:Ag assay, the vWF:CBA and the PFA-100; however, a corresponding pattern was not visually apparent in the raw data, making the association clinically irrelevant. The association between the platelet count and the PTT with the other additional tests was negligible. Based on our results, the vWF:CBA and PFA-100 would be valuable assets, in conjunction with a vWF:Ag assay, in a canine vWD diagnostic profile to further characterize patients with this disease.

The objective of this study was to use a questionnaire 1) for characterization of hemorrhagic signs; 2) to assess its value as a predictor of von Willebrand Disease (vWD) status; and 3) for evaluation of the vWD diagnostic profile [platelet function analysis using the PFA-100, Collagen binding assay (vWF:CBA), and vWF antigen ELISA (vWF:Ag)], partial thromboplastin time (PTT) and Factor VIII activity (FVIII) as predictors of hemorrhagic risk. von Willebrand factor (vWF) concentration and function was assessed for each of the 165 canine participants using the vWD diagnostic profile. Hemorrhagic signs for each dog were obtained using a standardized questionnaire. Questionnaires were scored according to a previously prepared scoring key. Of the 165 dogs in the study, 43.6% had a low vWF concentration, with only 48.6% of dogs in this group having reports of hemorrhagic signs. Oral bleeding was the most commonly reported sign. The questionnaire had a sensitivity of 48.6% and a specificity of 78.5% for the prediction of vWD status. Using the Spearman correlation coefficient, a statistical association was found between the questionnaire and the vWD diagnostic profile components. However, this could not be translated into an ability to predict hemorrhage. The questionnaire allowed characterization of hemorrhagic signs in a large population of dogs over a range of vWF:Ag concentrations, and demonstrated that the vWD diagnostic profile was unsuccessful in the prediction of hemorrhagic risk. Although the sensitivity was insufficient for a screening tool, the questionnaire did have some discriminatory power in the prediction of vWD status.

Background: Cardiac disease has the potential to alter platelet function in dogs. Evaluation of platelet function using the PFA-100 analyzer in dogs of multiple breeds and with a broad range of cardiac conditions would help clarify the effect of cardiac disease on platelets., Objectives: The objective of this study was to assess differences in closure time (CT) in dogs with cardiac disease associated with murmurs, when compared with that of healthy dogs., Methods: Thirty-nine dogs with cardiac murmurs and turbulent blood flow as determined echocardiographically were included in the study. The dogs represented 23 different breeds. Dogs with murmurs were further divided into those with atrioventricular valvular insufficiency (n=23) and subaortic stenosis (n=9). Fifty-eight clinically healthy dogs were used as controls. CTs were determined in duplicate on a PFA-100 analyzer using collagen/ADP cartridges., Results: Compared with CTs in the control group (mean+/-SD, 57.6+/-5.9 seconds; median, 56.5 seconds; reference interval, 48.0-77.0 seconds), dogs with valvular insufficiency (mean+/-SD, 81.9+/-26.3 seconds; median, 78.0 seconds; range, 52.5-187 seconds), subaortic stenosis (71.4+/-16.5 seconds; median, 66.0 seconds; range, 51.5-95.0 seconds), and all dogs with murmurs combined (79.6+/-24.1 seconds; median, 74.0 seconds; range, 48.0-187 seconds) had significantly prolonged CTs (P<.01)., Conclusions: The PFA-100 analyzer is useful in detecting platelet function defects in dogs with cardiac murmurs, most notably those caused by mitral and/or tricuspid valvular insufficiency or subaortic stenosis. The form of turbulent blood flow does not appear to be an important factor in platelet hypofunction in these forms of cardiac disease.

Background: Clinical diagnosis of platelet dysfunction is complex and technically challenging. The wide repertoire of platelet responses requires a test panel to assess different parameters of platelet reactivity. While "global" hemostasis analyzers and whole blood assays have potential for testing platelet function, their ability to evaluate platelet procoagulant activity is ill-defined., Objectives: The aim of this study was to determine whether platelet procoagulant deficiency, the pathophysiologic defect of Scott syndrome, could be detected in point-of-care and whole blood assays., Methods: Study subjects were 4 Scott syndrome-affected German Shepherds and 8 control dogs. We evaluated 2 point-of-care instruments: the platelet function analyzer (PFA-100) and thromboelastograph (TEG). TEG analysis was performed on recalcified citrated whole blood with and without tissue-factor activation. A whole blood flow cytometric assay was configured to detect thrombin-induced platelet P-selectin expression and platelet-derived microparticle release. Cytometric samples were analyzed after 1 hour and 1 day of storage., Results: We found no significant differences between Scott and control dogs in PFA-100 COL/ADP closure times or in any TEG parameter in tissue-factor-activated samples. In nonactivated samples, mean clotting time (K) and time to maximal rate of thrombus generation were significantly prolonged in Scott dogs; however, values overlapped with those of control dogs. Cytometric analysis of samples from Scott dogs showed significantly diminished platelet-derived microparticle release. Samples from all dogs reanalyzed after 1 day of storage had nonspecific increases in basal P-selectin expression and vesiculation., Conclusion: A whole blood cytometric assay to detect stimulated platelet microparticle release can be used to screen for Scott syndrome. However, platelet activation artifacts preclude overnight storage for next-day analysis.

The aim of this study was to evaluate platelet function in Dachshunds during early stages of myxomatous mitral valve disease. Clinical examination and echocardiography were performed in 34 wirehaired standard sized Dachshunds. Platelet function was evaluated using the PFA-100 (reported as closure time). In addition, whole blood platelet aggregation response and hemostatic markers were evaluated. Significant longer PFA-100 closure time (CT) was found in 12 Dachshunds with mild mitral regurgitation (MR) compared to 22 Dachshunds with minimal MR. Only five Dachshunds responded to adenosine diphosphate in the whole blood aggregation analyses. There were no differences between the two dog groups in plasma fibrinogen, plasma von Willebrand factor (vWf) or vWf multimer distribution; however, there was a significant correlation between CT and plasma vWf concentration and CT and plasma fibrinogen concentration. The higher CT found in Dachshunds with mild MR suggests a form of platelet dysfunction in Dachshunds with MR.

Objectives: To compare coagulation and platelet function parameters measured using a viscoelastic analyzer in 3 groups: foals presenting to a neonatal intensive care unit with presumed sepsis, normal foals, and adult horses., Design: Preliminary prospective trial., Setting: Veterinary teaching hospital., Animals: Ten clinically healthy foals, 13 clinically healthy adult horses, and 17 foals sequentially admitted for suspected sepsis. Intervention- A single citrated (3.8%) blood sample collected at admission was submitted for coagulation evaluation using a viscoelastic analyzer., Measurements and Main Results: Time to initial clot formation (ACT), clot rate (CR), platelet function, and time to peak parameters were collected from the signature generated with the associated software. Peak clot strength was collected manually from signature tracings. Signalment, presenting complaint, blood culture results, clinical progression, and outcome were collected from the medical record. Kruskal-Wallis testing was used to determine differences in coagulation parameters between groups, as well as to identify any associations between coagulation variables, foal variables, and outcome. Normal foals were more likely to have increased platelet function (P=0.04) compared with normal adult horses. Prolonged ACT (P=0.004) and decreased CR (P=0.03) were associated with foals with positive blood culture. There was a trend toward prolonged ACT and increased likelihood of death (P=0.06)., Conclusions: Healthy foals differ in values measured by the viscoelastic coagulation and platelet function analyzer compared with healthy adult horses. ACT and CR abnormalities were more likely to be observed in foals with positive blood cultures. The viscoelastic coagulation and platelet function analyzer may be useful in identifying early hemostasic and platelet dysfunction in critically ill foals, particularly those that are septic.

Background: There is currently no simple analytical tool for the evaluation of hypercoagulability in cats. The Platelet Function Analyzer-100 (PFA-100; Dade Behring Inc., Deerfield, IL, USA) is a bench-top machine that evaluates platelet function by measuring closure time (CT) in citrated whole blood under high shear conditions. We hypothesized that cats with hypertrophic cardiomyopathy (HCM) have up-regulated platelet function, which shortens their CT and increases their risk for thromboembolic events., Objectives: The goals of this study were to: (1) establish a feline reference interval for CT using the PFA-100, (2) measure CT in blood from cats with HCM, and (3) determine if there is a measurable difference between the CT of healthy cats compared with cats with HCM., Methods: Citrated blood samples from 42 clinically healthy cats and 30 cats with HCM were analyzed according to manufacturer's specifications. CT was measured in triplicate and the mean value was used for analysis. Transformed data were compared between clinically healthy cats and cats with HCM using a Student's t-test, and among cats with mild, moderate, or severe HCM using ANOVA., Results: The median CT of clinically healthy cats was 64 seconds (range 43-176 seconds). The median CT of cats with HCM was 74 seconds (range 48-197 seconds). There was no significant difference in CT between cats with HCM and clinically healthy cats. There also were no significant differences in cats with mild, moderate, or severe HCM., Conclusions: A feline reference interval for PFA-100 CT will be useful in future studies of platelet function in cats. Cats with HCM do not have shorter CTs when compared with clinically healthy cats.

Functional defects of platelets are often studied by in-vitro aggregation tests with chemical compounds such as ADP, epinephrine and ristomycin (ristocetin). The aim of the present work was to investigate the effect of some diseases and that of nonsteroidal anti-inflammatory drug treatment on platelet aggregation in dogs. The examination had been carried out on 115 dogs by a Carat TX4 optical aggregometer (Entec GmbH, Ilmenau, Germany) first used in veterinary practice. The dogs were divided in three groups: healthy (control) dogs (n = 43), diseased dogs with normal haemostasis profile (n = 44), and dogs suffering from arthropathies with normal haemostasis profile treated with the nonsteroidal anti-inflammatory drugs ketoprofen or carprofen (n = 21). Following establishment of normal platelet aggregation curves in healthy dogs we found that some diseases such as diabetes mellitus, Cushing's disease, mastocytoma and lymphoma increased or decreased the aggregation maximum of platelets or caused changes in the feature of the aggregation curve. Carprofen treatment had no effect on platelet aggregation while ketoprofen decreased the aggregation maximum. These results showed that Carat TX4 aggregometer proved a useful instrument in studying platelet aggregation in platelet-rich plasma of dogs. For clinical pathologists it is important to know that the effects of some diseases and nonsteroidal anti-inflammatory drug treatments have to be taken into account when in-vitro platelet aggregation is evaluated. Based on our results and on those of other studies, we think standardization in aggregation methodology is highly recommended in veterinary laboratories.

The effect of two nonsteroidal anti-inflammatory drugs (carprofen and ketoprofen) on platelet adhesion and aggregation functions was evaluated by the PFA-100 analyser (Dade-Behring, CA, U.S.A.) using its collagen-adenosine diphosphate (ADP) and collagen-epinephrine (EPI) cartridges. The function of platelets was evaluated in 55 healthy dogs, in 7 dogs treated with ketoprofen and in 31 dogs treated with carprofen in a therapeutic dose for minimum 5 days. The therapeutic doses of carprofen had no effect on the closure time of PFA-100 (which is the marker of platelet function) but ketoprofen caused a significant increase when using collagen-EPI stimulation The closure times for both the healthy (control) and the treated dogs using EPI cartridges were often longer than the upper default cut-off point (300 sec) of the device. The PFA-100 analyser with collagen-ADP cartridges could be a useful tool for veterinary applications including the evaluation of platelet aggregation in dogs treated with NSAIDs. The upper cut-off point of PFA-100 might be extended.

Background: Blood collection tubes containing 3.2% (0.109 M) sodium citrate, instead of 3.8% (0.129 M) sodium citrate, have recently become available in the United States. These tubes are visually indistinguishable from the traditional 3.8% sodium citrate tubes, except for wording on the label. Consequently, samples for hemostatic evaluation are frequently collected in tubes containing the lower concentration of sodium citrate., Hypothesis: Results of hemostasis assays are different in samples collected in 3.2% versus 3.8% sodium citrate., Animals: Twenty healthy dogs., Methods: This study aimed at determining whether results of standard coagulation tests, von Willebrand factor concentration (vWF:Ag), and platelet function with the platelet function analyzer PFA-100a were affected by the different concentrations of sodium citrate. Blood samples were collected in tubes containing either 3.2% or 3.8% sodium citrate concentrations and processed routinely for coagulation assays (one-stage prothrombin time [OSPT], activated partial thromboplastin time [aPTT], fibrinogen concentration, and platelet count), vWF:Ag, and platelet function assays with a PFA-100., Results: There was no significant difference between samples collected in 3.2% versus those collected in 3.8% sodium citrate for OSPT, aPTT, fibrinogen concentration, platelet count, or vWF:Ag. The closure times with collagen/adenosine diphosphate were significantly shorter (66 +/- 8.1 versus 74.8 +/- 9.7 seconds; P < .0001) with the 3.2% than with 3.8% sodium citrate concentration, and the hematocrit was significantly higher (47.9 +/- 5.6 versus 46.0 +/- 4.7 seconds; P = .03) in samples collected in 3.2% than in those collected in 3.8% sodium citrate., Conclusions and Clinical Importance: There is no clinically relevant effect of collection of blood into 3.2% or 3.8% sodium citrate.

Haemostatic alterations in dogs naturally infected by ehrlichiosis and/or leishmaniasis were studied. Platelet count, ADP and collagen-induced platelet aggregation, prothrombin time, activated partial thromboplastin time (APTT) and plasma fibrinogen concentration were measured. An evident reduction of platelet aggregation response was shown for Leishmania-Ehrlichia co-infected dogs where platelet aggregation was lower in comparison with control and leishmaniotic dogs (ADP and collagen, P < or = 0.01) and ehrlichiotic dogs (ADP 10 and 7.5 microm, P < or = 0.05). Moreover, a significant increase in APTT as well as a reduction of the albumin/globulin rate (A/G) for leishmaniotic and co-infected dogs versus control and ehrlichiotic dogs was detected. The hypothesis of a synergism between leishmaniosis and ehrlichiosis in altering platelet function by different pathways is discussed.

Veterinarians involved in Greyhound rescue have anecdotally observed that 10-15% of Greyhounds bleed profusely after simple surgical procedures. In most patients, platelet counts and hemostasis profiles are normal; therefore, it is possible that these dogs have platelet dysfunction. The PFA-100 is a novel point-of-care platelet function analyzer that has recently been evaluated as a rapid method to assess platelet function in dogs. The objectives of this study were to characterize platelet function in a group of healthy Greyhounds by means of the PFA-100. Blood samples were collected from the jugular vein from 30 healthy Greyhounds. CBC, biochemical profile, PFA-100 assay with collagen/epinephrine (COL-EPI) and collagen/ adenosindiphosphate (COL-ADP), plasma von Willebrand factor antigen concentration (vWF:Ag), and vWF collagen-binding assay (vWF:CBA) were performed. PFA-100 closure times (CTs) with COL/ADP ranged from 63 to 92 seconds (mean +/- SD, 74.7 +/- 7.9 seconds) and with COL/EPI from 87 to 238 seconds (138 +/- 41 seconds); vWF: Ag ranged from 22 to 120% (87.52 +/- 25.5%) and vWF: CBA ranged from 36 to 102% (77.4 +/- 17.3%); and platelet counts ranged from 147 to 265 x 10(9)/L (194.6 +/- 31.64 x 10(9)/L). Greyhound CTs were significantly shorter than CTs in a mixed population of 50 healthy non-Greyhound dogs, in which the COL/ADP CTs ranged from 61 to 172 seconds (mean +/- SD, 87 +/- 21.6 seconds), and the COL/ EPI CTs ranged from 81 to 300 seconds (mean +/- SD, 183 +/- 67.6 seconds; P = 0.005 for COL/ADP CT; P = 0.001 for COL/ EPI CT). Also, platelet counts were significantly lower (P = 0.001) and packed cell volume was significantly higher (P = 0.001) in the Greyhound than in the non-Greyhound group. The PFA-100 is a reproducible method that can be used in the clinical setting to assess platelet function in Greyhounds; however, normal CTs in healthy Greyhounds are shorter than in other breeds. The results obtained in this study will be used to screen for abnormal platelet function in Greyhounds with postoperative bleeding.

We report here on the influence of the platelet antagonist clopidogrel (Plavix) on bovine platelet function. We first evaluated the capacity of clopidogrel to inhibit adenosine diphosphate (ADP)-stimulated platelet function in the bovine species, using an ex vivo approach with blood from treated animals. Platelets isolated from treated calves displayed rapid and consistent reduction in function (aggregation, thromboxane production) upon ADP, but not platelet activating factor (PAF), stimulation. We then examined the possibility that clopidogrel could influence Mannheimia haemolytica pneumonia pathobiology using an experimental challenge model. We were unable to detect significant differences between clopidogrel treated and untreated animals when challenged with intra-tracheal inoculation of M. haemolytica. There was a trend towards inhibition of platelet degranulation in the affected regions of lungs from clopidogrel treated calves, and pre-treated challenged animals had similar amounts of fibrin deposition and enhanced fibrous tissue formation in their lungs when compared with control counterparts.

The objective of this study was to assess whether a new human platelet function analyser (the PFA-100) could be used to evaluate platelet function in horses and detect acetylsalicylic acid (ASA)-induced platelet dysfunctions. Citrated blood samples from 40 healthy horses were processed to obtain reference values for closure time (CT) using cartridges with collagen-ADP (CT-ADP) and collagen-epinephrine (CT-EPI) as platelet agonists. In addition, CT-ADP and CT-EPI were also measured before and 24 h after oral ASA administration in another 12 healthy horses. The sensitivity and specificity of the test were also determined. In normal horses, means+/-SD value for CT-ADP was 85.1+/-13.1 s (median, 82 s), and CT-EPI ranged from 158 to >300 s (median 291 s). Calculated reference ranges were 60.5-115.9 s and 158.5->300 s for CT-ADP and CT-EPI, respectively. Administration of ASA significantly (P<0.001) prolonged CT-ADP values from 91.0+/-13 to 113.5+/-14.4 s, and CT-EPI values were also significantly (P<0.008) prolonged after ASA administration. Sensitivity and specificity results for ADP cartridges showed that a prolonged CT value would be highly suggestive of a platelet aggregation inhibition. In conclusion, ADP cartridges can be used in horses to assess primary haemostasis and may be a valuable test for the detection of platelet aggregation inhibition.

With aggregometry, increased platelet activity has been reported in Cavalier King Charles Spaniels (CKCS) without mitral regurgitation (MR). In contrast, dogs with MR have been found to have decreased platelet activity. The purpose of this study was to test an easy bedside test of platelet function (the Platelet Function Analyzer [PFA-100]) to see if it could detect an increase in platelet activity in CKCS without MR and a decrease in platelet activity in CKCS with MR. This study included 101 clinically healthy dogs > 1 year of age: 15 control dogs of different breeds and 86 CKCS. None of the dogs received medication or had a history of bleeding. The PFA-100 evaluates platelet function in anticoagulated whole blood under high shear stress. Results are given as closure times (CT): the time it takes before a platelet plug occludes a hole in a membrane coated by agonists. The CT with collagen and adenosine-diphosphate as agonists was similar in control dogs (median 62 seconds; interquartile interval 55-66 seconds) and CKCS with no or minimal MR (55; 52-64 seconds). The CT was higher in CKCS with mild MR (regurgitant jet occupying 15-50% of the left atrial area) (75; 60-84 seconds; P = .0007) and in CKCS with moderate to severe MR (jet > 50%) (87: 66-102 seconds; P < .0001). CKCS with mild, moderate, and severe, clinically inapparent MR have decreased platelet function. The previous finding of increased platelet reactivity in nonthrombocytopenic CKCS without MR could not be reproduced with the PFA-100 device.

The platelet function analyser PFA-100 aspirates blood in vitro from a sample reservoir in disposable test cartridges through a microscopic aperture cut into a biologically active membrane at the end of a capillary. In different cartridges the membrane is coated with collagen and adenosine diphosphate (ADP) or collagen and epinephrine (adrenaline) inducing a platelet plug and closure of the aperture. The closure time and total volume of blood flow through the capillary until closure of its aperture were measured. The correlation between platelet count in samples of thrombocytopenic dogs and results of the collagen/ADP cartridge (closure time: r(S)=-0.579; total volume: r(S)=-0.549) was closer than between platelet count and capillary bleeding time. No significant correlation was observed between platelet count and the results obtained with the collagen/epinephrine cartridge. In addition, a higher sensitivity was obtained for the collagen/ADP cartridge. Injection of acetylsalicylic acid into healthy dogs significantly increased closure time and total volume of both types of cartridges (P<0.01). Two dogs with von Willebrand's disease had abnormal values. In contrast, coagulopathies did not significantly influence the results of the platelet function analyser (P>0.05). Despite adequate sensitivity of measurements using the collagen/ADP cartridge to assess quantitative and qualitative platelet disorders in dogs, the influence of haematocrit (P<0.0001) will limit the clinical application of the analyser.

Hereditary platelet function disorders are clinically characterized by recurrent surface bleeding and prolonged bleeding time, despite normal platelet count and coagulation tests. The authors describe persistent thrombopathies in two young dogs with increased bleeding tendencies but with normal plasma coagulation times and von Willebrand factor (vWf) concentrations. Buccal mucosal bleeding times were prolonged in both dogs. In aggregation studies, platelets underwent only a shape change or minimal aggregation in response to adenosine diphosphate and collagen. Whole-platelet adenine nucleotide concentrations were normal. Electron microscopic evaluation of fibrinogen and vWf binding to the platelets of case no. 1 demonstrated the presence of glycoprotein IIb/IIIa and Ib receptors. Thus, the intrinsic platelet function defects may be different in these two dogs and may likely represent secretion/signal transduction disorders.

Platelet function was assessed by aggregometry in 10 Holstein calves before and after exposure to Pasteurella haemolytica (biotype A, serotype 1) by intrabronchial challenge. At 24 h after exposure the platelets had become more reactive to stimulation with known platelet agonists such as adenosine diphosphate (ADP) and platelet-activating factor (PAF) and the platelet aggregates that formed were more resistant to disaggregation. The activation of platelets was an early response in the challenged calves as platelet function had returned to pretreatment levels 72 h after exposure to the bacteria while the acute phase reactant proteins, haptoglobin and fibrinogen, were approaching their peak values and alpha 2-macroglobulin levels had also risen significantly (P < 0.05) at this time. The plasma levels of these proteins were still elevated and albumin levels were depressed 6 d post-treatment. At post-mortem all calves exhibited pneumonic tissue damage. When P. haemolytica leukotoxin was added directly to bovine platelet suspensions both spontaneous aggregation and an increase in the aggregation response to ADP and PAF stimulation were observed. The morphological appearance of the platelet aggregates exhibited the typical pattern for bovine platelets with 2 distinct zones of cells being visible within each aggregate. One zone contained platelets in which the cytoplasmic granules were still evident and the other zone contained irregularly shaped platelets devoid of granular content. In the latter zone, discrete gaps, or pores, were evident in the plasma membrane of numerous platelets. This pore formation is characteristic of leukotoxin action and is not observed in ADP or PAF induced aggregates.

Objective: To establish the existence of platelet-derived proteins in equine plasma, with the future goal of developing an assay for the detection of in vivo platelet activation., Animals: 5 mature healthy horses., Procedure: Platelet-rich plasma and platelet-poor plasma were prepared from anticoagulated blood. Platelets were separated from plasma proteins by gel filtration, then activated with 0.5 microM platelet-activating factor. Protease inhibitors were added, and the released platelet proteins were harvested. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis was performed on the released platelet proteins and platelet-poor plasma, and the resultant silver-stained bands were compared. Immunoblot analysis was performed on released platelet proteins, using an antibody to human thrombospondin; human platelet-derived proteins served as the positive control for the antibody., Results: Released platelet proteins in the presence of beta-mercaptoethanol (reduced samples) contained several proteins that were not observed in plasma including (mean +/- SEM) 194 +/- 2, 159 +/- 2, 151 +/- 2, 104 +/- 2, and 95 +/- 1 kd. Immunoblots of released platelet proteins had a prominent 180 +/- 2-kd protein in reduced samples that was recognized by an antibody to human thrombospondin, and with prolonged color development, 2 additional less prominent proteins (166 +/- 1 and 155 +/- 1 kd) were observed., Conclusions: Several proteins are released from activated equine platelets that are not detectable in normal equine plasma. Thrombospondin is one of the high molecular mass proteins released by activated equine platelets., Clinical Relevance: An assay can be developed for detection of thrombospondin in equine plasma and may be useful for detection of in vivo platelet activation in horses.

A severe bleeding disorder in Simmental cattle has been described in widespread locations in the USA and Canada. The clinical findings are consistent with a hemophilia-like disease or, more precisely, a hereditary hemorrhagic diathesis and include spontaneous epistaxis, hematuria, and excessive bleeding associated with trauma or standard management procedures such as tattooing, ear tagging, and castration. A preliminary investigation of this defect showed that blood-platelet numbers and coagulation profiles of affected cattle were normal. Affected animals have a marked dysfunction of platelets (thrombopathy), termed Simmental hereditary thrombopathy. The defect is very similar or identical to that described in the same breed by 2 other laboratories.

A bleeding disorder due to abnormal platelet function occurs in Simmental cattle. Whole blood from these animals underwent good clot retraction. Platelet aggregation in response to adenosine diphosphate (ADP) and collagen in a whole blood aggregation system was markedly impaired. Normal bovine platelets in a whole blood aggregation system showed very little aggregation in response to epinephrine and arachidonic acid. Aggregation in platelet-rich plasma was negligible in response to ADP, collagen and thrombin. Dense granule release of radiolabelled serotonin from the platelets of one affected cow was similar to that of normal bovine platelets. Platelet membrane glycoprotein electrophoresis with the platelets of one affected cow revealed no quantitative abnormalities. These findings reveal similarities and differences in thrombopathic Simmental platelet function when compared to human Glanzmann's thrombasthenia and Basset Hound thrombopathia.

Blood obtained from Mongolian gerbils (Meriones unguiculatus) by cardiac puncture was used to develop a method for preparing platelet suspensions suitable for biochemical and aggregometric studies. The aggregation responses of gerbil platelet suspensions to 10 microM adenosine diphosphate and to 0.2 U/mL thrombin were immediate and irreversible. In addition, thrombin produced a short lag period. Bovine acid-soluble collagen (1:800) produced a long lag period coupled with an irreversible aggregation. Indomethacin (10 microM) significantly depressed adenosine diphosphate--induced aggregation but had no effect on thrombin-induced aggregation. Indomethacin significantly prolonged the lag period and reduced the rate of collagen-induced aggregation. Our results strongly indicate that platelet suspensions from the Mongolian gerbil, an animal which is responsive to hypercholesterolemic diets, may be useful in studying dietary lipid factors which influence platelet aggregation.

The diagnosis of hemostatic abnormalities requires a detailed clinical and laboratory evaluation of the patient. The clinical assessment includes a detailed history and a thorough physical examination. The patient's history may provide clues as to the time of onset of the bleeding tendency, the clinical severity of the abnormality, and the possible contributing role of other disease processes or of drugs. The nature of the bleeding symptoms may provide clues as to the nature of the hemostatic defect. Hemostatic screening tests are invaluable in helping to differentiate between platelet, vascular, coagulation, and fibrinolytic abnormalities. Specific tests, including specific factor assays, platelet aggregometry, and antiplatelet antibody assays are usually required to characterize the exact nature and severity of hemostatic defects.

Animals with disorders of hemostasis are often presented as emergency patients and, as such, offer a challenge to the attending clinician. This article reviews the basic physiology of hemostasis and laboratory tests used for diagnosis. Guidelines for the evaluation and treatment of patients with bleeding disorders are provided.

Thrombocytopenia may be caused by abnormal platelet production, accelerated removal owing to immunologic or nonimmunologic reasons, or sequestration of platelets in the spleen. Bleeding associated with thrombocytopenia usually presents as petechial or ecchymotic hemorrhages or epistaxis. Immunologic and nonimmunologic cases of thrombocytopenia may be diagnosed with routine hematology, bone marrow cytology, and platelet specific tests. Thrombocythemia may also be associated with platelet functional abnormalities, contrasting the normal platelet function noted in reactive thrombocytosis.

A prolonged mean template bleeding time of 13 minutes was present in nine Aleutian mink affected with Chediak-Higashi syndrome (CHS) compared with 4 minutes in dark control mink. The concentrations of blood platelets in normal and affected animals did not differ significantly. However, in mink with CHS, a marked disturbance of platelet response to collagen was present. Administration of aspirin and indomethacin completely blocked CH platelet response to collagen. Blood platelet adenosine triphosphate and adenosine diphosphate values from mink with CHS were significantly less than those of normal mink, and the platelet adenosine triphosphate/adenosine diphosphate ratios were 10.31 in affected mink and 2.74 in normal mink. These findings are consistent with our previous investigations in affectd cattle and persons and indicate that a "storage pool disease" of platelets exist in the mink with CHS.

In this paper we describe our clinical experience and results with the cuticle bleeding time test in a colony of cross-bred Labrador retrievers with severe haemophilia A. The dogs have a severe bleeding tendency with a high incidence of fatal haemorrhages in the central nervous system. Homozygous females appeared to be especially prone to this lethal complication. Factor VIII recovery and half-life determinations yielded results similar to the data from human studies. The cuticle bleeding time proved to be a good measure of the coagulation defect. The prolongation of the bleeding time could be completely abolished by administration of 10 to 15 units of canine factor VIII per kg body weight. We conclude that the cuticle bleeding time in canine haemophilia provides us with a suitable model for the in vivo study of new therapeutic materials.

Changes in the production of proaggregatory (thromboxane A2 and prostaglandin E2) and antiaggregatory (prostacyclin) prostaglandins by blood platelets, macrophages and endothelial cells during acute African swine fever caused by both a highly virulent virus and a less virulent virus were studied. No impairment in thromboxane A2 release by either platelets or macrophages could be detected but prostacyclin production by the endothelium was impaired. There was also a significant increase in prostaglandin E2 release by macrophages at the time when thrombocytopenia was most marked. However, the early event that causes primary aggregation remains obscure.