Summary: Introduction: The investigation of platelet function by aggregometry requires specialist equipment and is labour intensive. We have developed an automated platelet aggregation method on a routine coagulation analyser.Methods: We used a CS‐2000i (Sysmex) with prototype software to perform aggregation in platelet‐rich plasma (PRP), using the following agonists: ADP (0.5–10 μm), epinephrine (0.5–10 μm), collagen (0.5–10 mg/μL), ristocetin (0.75–1.25 mg/mL) and arachidonic acid (0.12–1.0 mm). Platelet agonists were from Hyphen Biomed, and an AggRAM aggregometer (Helena Biosciences) was used as the reference instrument.Results: CS‐2000i reaction cuvette stirrer speed was found to influence reaction sensitivity and was optimized to 800 rpm. There were no clinically significant changes in aggregation response when the PRP platelet count was 150–480 x 10⁹/L, but below this there were changes in the maximum amplitude (MA) and slope (rate). Dose response with each of the agonists was comparable between CS‐2000i and an AggRAM aggregometer and normal subjects receiving antiplatelet drugs. Aggregation imprecision was similar on both the CS‐2000i and AggRAM systems, with a cv for 2–5 μm ADP MA and slope varying between 3–12%.Conclusion: Our preliminary studies indicated that optimal sensitivity using the CS‐2000i was obtained with a reaction cuvette stirrer speed of 800 rpm and a PRP platelet count of 200–300 x 10⁹/L; aggregation with a PRP count <100 x 10⁹/L showed poor sensitivity. Imprecision and detection of antiplatelet drug effects was similar between the CS‐2000i and AggRAM. These data demonstrate that CS‐2000i is comparable to a stand‐alone aggregometer, although CS‐2000i has the advantages of walk‐away technology and also required a smaller sample volume than the AggRAM (44% less).

Introduction: Plasma samples with gross lipaemia present a challenge for coagulation laboratories using optical analysers. High‐speed centrifugation may be used to remove excess lipids but it has not established whether this affects haemostasis tests. The aims were to determine whether the removal of lipid by centrifugation affects PT, APTT, fibrinogen, D‐dimer and von Willebrand factor activity measurements.Methods: Twenty‐six lipaemic samples (median [range]): triglyceride 4.6 mmol/L [0.5‐17.0]; cholesterol: 4.06 mmol/L [2.20‐9.41] and 20 plasmas spiked with Intralipid 20 or lipid isolated from patient plasmas (median triglyceride of 11.95 mmol/L [5.0‐17.0] and cholesterol 4.33 [3.22‐7.06]), were tested before and after the removal of the lipid layer by centrifugation (10000 g for 10 minutes). Tests were performed using the CS‐5100 (Sysmex) coagulation analyser.Results: Thirteen, 9, 3 and 1 of the lipaemic or spiked samples failed to give PT, APTT, fibrinogen and D‐dimer results, respectively. Centrifugation significantly reduced triglyceride (median 2.7, [0‐6.1 mmol/L]) and cholesterol (median 0.52 [0‐3.5]), allowing clot detection in all tests. There were no statistically significant differences in fibrinogen, D‐dimer or VWF levels in samples before and after lipid removal. A small but clinically insignificant change in PT and APTT was observed after lipid removal.Conclusion: High‐speed centrifugation reduces lipaemia sufficiently to allow testing on an optical coagulation analyser without introducing clinically significant differences PT, APTT, fibrinogen, D‐dimer or VWF activity values.

Background Variability remains a challenge in lupus anticoagulant (LA) testing. Objective To validate LA test performance between Antiphospholipid Syndrome Alliance for Clinical Trials and International Networking (APS ACTION) Core laboratories and examine agreement in LA status between Core and local/hospital laboratories contributing patients to this prospective registry. Methods Five Core laboratories used the same reagents, analyzer type, protocols, and characterized samples for LA validation. Non-anticoagulated registry samples were retested at the corresponding regional Core laboratories and anticoagulated samples at a single Core laboratory. Categorical agreement and discrepancies in LA status between Core and local/hospital laboratories were analyzed. Results Clotting times for the reference/characterized plasmas used for normalized ratios were similar between Core laboratories (CV <4%); precision and agreement for LA positive/negative plasma were similar (all CV ≤5%) in the four laboratories that completed both parts of the validation exercise; 418 registry samples underwent LA testing. Agreement for LA positive/negative status between Core and local/hospital laboratories was observed in 87% (115/132) non-anticoagulated and 77% (183/237) anticoagulated samples. However, 28.7% (120/418) of samples showed discordance between the Core and local/hospital laboratories or equivocal LA results. Some of the results of the local/hospital laboratories might have been unreliable in 24.7% (41/166) and 23% (58/252) of the total non-anticoagulated and anticoagulated samples, respectively. Equivocal results by the Core laboratory might have also contributed to discordance. Conclusions Laboratories can achieve good agreement in LA performance by use of the same reagents, analyzer type, and protocols. The standardized Core laboratory results underpin accurate interpretation of APS ACTION clinical data.

Background: Acquired activated protein C resistance (APCr) has been identified in antiphospholipid syndrome (APS) and systemic lupus erythematosus (SLE). Objective: To assess agreement between the ST-Genesia^® and CAT analysers in identifying APCr prevalence in APS/SLE patients, using three thrombin generation (TG) methods. Methods: APCr was assessed with the ST-Genesia using STG-ThromboScreen and with the CAT using recombinant human activated protein C and Protac^® in 105 APS, 53 SLE patients and 36 thrombotic controls. Agreement was expressed in % and by Cohen’s kappa coefficient. Results: APCr values were consistently lower with the ST-Genesia^® compared to the CAT, using either method, in both APS and SLE patients. Agreement between the two analysers in identifying APS and SLE patients with APCr was poor (≤65.9%, ≤0.20) or fair (≤68.5%, ≥0.29), regardless of TG method, respectively; no agreement was observed in thrombotic controls. APCr with both the ST Genesia and the CAT using Protac^®, but not the CAT using rhAPC, was significantly greater in triple antiphospholipid antibody (aPL) APS patients compared to double/single aPL patients (p < 0.04) and in thrombotic SLE patients compared to non-thrombotic SLE patients (p < 0.05). Notably, the ST-Genesia^®, unlike the CAT, with either method, identified significantly greater APCr in pregnancy morbidity (median, confidence intervals; 36.9%, 21.9–49.0%) compared to thrombotic (45.7%, 39.6–55.5%) APS patients (p = 0.03). Conclusion: Despite the broadly similar methodology used by CAT and ST-Genesia^®, agreement in APCr was poor/fair, with results not being interchangeable. This may reflect differences in the TG method, use of different reagents, and analyser data handling.