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Analytical validation of a flow cytometric protocol for quantification of platelet microparticles in dogs

Signe E Cremer1, Anne K H Krogh1, Matilda E K Hedström1

  • 1Department of Veterinary and Animal Sciences, University of Copenhagen, Copenhagen, Denmark.

Veterinary Clinical Pathology
|March 31, 2018
PubMed
Abstract

Insights

This study validates a canine flow cytometry protocol for quantifying platelet microparticles (PMPs). Reliable PMP quantification in whole blood shows potential for clinical use, unlike unreliable plasma analysis.

Area of Science:

  • Veterinary Hematology
  • Flow Cytometry
  • Platelet Biology

Background:

  • Platelet microparticles (PMPs) are vesicles released during platelet activation.
  • PMP alterations are linked to human diseases, but canine studies are limited.

Purpose of the Study:

  • To validate a canine flow cytometry protocol for PMP quantification.
  • To assess the impact of calcium on PMP concentrations in dogs.

Main Methods:

  • Flow cytometry was used to quantify microparticles (MPs) in canine whole blood and plasma.
  • Anti-CD61 and Annexin V antibodies identified platelet-derived MPs and phosphatidylserine exposure.
  • Intra-assay and inter-assay coefficients of variation (CVs) and detection limits (DLs) were calculated.

Main Results:

  • Calcium buffer increased in vitro PMP concentrations (CD61+/AnV-).
  • Whole blood PMP quantification (CD61+/AnV- and CD61+/AnV+) showed acceptable intra- and inter-assay CVs in most dogs.
  • Platelet-poor plasma analysis yielded unreliable results due to high inter-assay CVs and interference from hemolysis/lipemia.

Conclusions:

  • Calcium enhances PMP concentrations in vitro, likely via platelet activation.
  • Canine whole blood PMP quantification is reliable and shows clinical potential.
  • Platelet-poor plasma analysis for PMPs in dogs is currently unreliable.
Keywords:
AnnexinCD61calciummicrovesiclesprecision

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