Analytical validation of platelet microparticle quantification in cats

BACKGROUND Cardiogenic embolism (CE) in cats is a devastating condition primarily associated with hypertrophic cardiomyopathy (HCM). Hypercoagulability may pose a risk for thrombus formation; however, no single test can predict CE development. Platelet microparticles (PMPs) released from platelet membranes are associated with thrombosis in humans. OBJECTIVES The aims were to validate flow cytometric PMP quantification in cats analytically and, in a pilot study, evaluate the procoagulant annexin V (AnV) positive PMP concentration in healthy cats and cats with asymptomatic HCM. METHODS With CD61 as a platelet marker, CD61+ AnV+ PMPs (0.3-1.0 μm) were quantified in citrated whole blood (WB) and platelet-poor plasma (PPP) using flow cytometry. Analyses were performed in 6 healthy cats and 5 cats with asymptomatic HCM. The coefficient of variation (CV) for duplicate (intra-assay) and parallel (inter-assay) analyses were calculated. RESULTS PMP concentrations were quantified with acceptable intra-assay CV for WB (CD61+ /AnV- ; 2.4%, 0.2%-8.4% (median, range), CD61+ /AnV+ ; 3.8%, 0.1%-12.5%) and PPP (CD61+ /AnV- ; 5.0%, 0.7%-12.8%, CD61+ /AnV+ ; 7.4%, 0.5%-15.3%), and acceptable inter-assay CV for WB in 10/11 cats (CD61+ /AnV- ; 6.2%, 1.4%-13.3%, CD61+ /AnV+ ; 6.4%, 0.7%-17.2%), but unacceptable for PPP (CD61+ /AnV- ; 15.6%, 5.8%-42.7%, CD61+ /AnV+ ; 27.8%, 8.4%-77.1%). For WB PMP concentrations, the pilot data demonstrated no differences between healthy cats and cats with asymptomatic HCM (4/5 with left ventricular outflow obstruction) for either the CD61+ /AnV- or the CD61+ /AnV+ PMPs. CONCLUSIONS Only WB PMP concentrations could be quantified reliably in cats in a clinical setting. PMP concentrations did not differ between healthy and asymptomatic HCM cats in this pilot study.