title: Whole blood flow cytometry measurements of in vivo platelet activation in critically-Ill patients are influenced by variability in blood sampling techniques.
abstract: BACKGROUND
  Flow cytometry is often used to measure in vivo platelet activation in critically-ill patients. Variability in blood sampling techniques, which may confound these measurements, remains poorly characterized.
  
  
  METHODS
  Platelet activation was measured by flow cytometry performed on arterial and venous blood from 116 critically-ill patients. We determined how variability in vascular sampling site, processing times, and platelet counts influenced levels of platelet-monocyte aggregates (PMA), PAC-1 binding (for glycoprotein (GP) IIbIIIa), and P-selectin (P-SEL) expression.
  
  
  RESULTS
  Levels of PMA, but not PAC-1 binding or P-SEL expression, were significantly affected by variability in vascular sampling site. Average PMA levels were approximately 60% higher in whole blood drawn from an arterial vessel compared to venous blood (16.2±1.8% vs. 10.7±1.2%, p<0.05). Levels of PMA in both arterial and venous blood increased significantly during ex vivo processing delays (1.7% increase for every 10 minute delay, p<0.05). In contrast, PAC-1 binding and P-SEL expression were unaffected by processing delays. Levels of PMA, but not PAC-1 binding or P-SEL expression, were correlated with platelet count quartiles (9.4±1.6% for the lowest quartile versus 15.4±1.6% for the highest quartile, p<0.05).
  
  
  CONCLUSIONS
  In critically-ill patients, variability in vascular sampling site, processing times, and platelet counts influence levels of PMA, but not PAC-1 binding or P-SEL expression. These data demonstrate the need for rigorous adherence to blood sampling protocols, particularly when levels of PMA, which are most sensitive to variations in blood collection, are measured for detection of in vivo platelet activation.
authors.affiliation: Divisions of General Internal Medicine, Department of Internal Medicine, Program in Molecular Medicine; University of Utah School of Medicine, Salt Lake City, Utah 84132, USA. matthew.rondina@hsc.utah.edu
authors.forename: Matthew T
authors.initials: MT
authors.lastname: Rondina
authors.forename: Colin K
authors.initials: CK
authors.lastname: Grissom
authors.forename: Shaohua
authors.initials: S
authors.lastname: Men
authors.forename: Estelle S
authors.initials: ES
authors.lastname: Harris
authors.forename: Hansjorg
authors.initials: H
authors.lastname: Schwertz
authors.forename: Guy A
authors.initials: GA
authors.lastname: Zimmerman
authors.forename: Andrew S
authors.initials: AS
authors.lastname: Weyrich
mesh_terms: D015415:Biomarkers / Q000097:blood; D001792:Blood Platelets / Q000502:physiology*; D001800:Blood Specimen Collection / Q000379:methods*; D016638:Critical Illness; D005260:Female; D005434:Flow Cytometry / Q000379:methods*; D006801:Humans; D008297:Male; D008875:Middle Aged; D018962:Phlebotomy; D015539:Platelet Activation / Q000502:physiology*
chemical_list: D015415:Biomarkers