Document Detail

Standardization of platelet-derived microparticle counting using calibrated beads and a Cytomics FC500 routine flow cytometer: a first step towards multicenter studies?
MedLine Citation:
PMID:  18983485     Owner:  NLM     Status:  MEDLINE    
BACKGROUND: Platelet microparticles (PMPs) have proved useful to identify patients with vascular risk. However, PMP counting, which is currently done by flow cytometry (FCM), needs to be standardized. OBJECTIVES: The objectives were (i) to standardize FCM settings for PMP counts on a routine instrument (Cytomics FC500) using size-calibrated fluorescent beads; (ii) to determine intra-instrument and inter-instrument reproducibility; and (iii) to establish PMP values in healthy subjects. METHODS: Using a blend of size-calibrated fluorescent beads (0.5 and 0.9 mum) in a fixed numerical ratio (Megamix), we gated PMPs in a restricted size window. To test intra-instrument and inter-instrument reproducibility, annexin V and CD41 coexpression were used to count PMPs in frozen aliquots of the same platelet-free plasma (PFP) over 4 months and in PFP from 10 healthy subjects on three independent flow cytometers. RESULTS: This calibrated-bead strategy allowed full long-term control of the FCM-based microparticle protocol and reproducible PMP counts over time [coefficient of variation (CV) < 10%]. Optimal settings were easily transferred from one instrument to another, using Megamix as a stable template. Similar PMP counts (CV < 12%) were obtained using the three instruments. With such a standardized FCM protocol, PMP values were established in healthy subjects (n = 60) with significantly higher levels in women than in men [median (1st quartile to 3rd quartile): 1775 microL(-1) (1014-3039 microL(-1)) vs. 656 microL(-1) (407-962 microL(-1))]. CONCLUSIONS: The present strategy provides a new option for PMP count standardization and thus opens the way for multicenter studies.
S Robert; P Poncelet; R Lacroix; L Arnaud; L Giraudo; A Hauchard; J Sampol; F Dignat-George
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Publication Detail:
Type:  Journal Article     Date:  2008-10-18
Journal Detail:
Title:  Journal of thrombosis and haemostasis : JTH     Volume:  7     ISSN:  1538-7836     ISO Abbreviation:  J. Thromb. Haemost.     Publication Date:  2009 Jan 
Date Detail:
Created Date:  2008-12-18     Completed Date:  2009-02-27     Revised Date:  -    
Medline Journal Info:
Nlm Unique ID:  101170508     Medline TA:  J Thromb Haemost     Country:  England    
Other Details:
Languages:  eng     Pagination:  190-7     Citation Subset:  IM    
Unité Mixte de Recherche S 608 (UMR-S 608), Institut National de la Santé et de la Recherche Médicale (INSERM), Université de la Méditerranée, Marseille, France.
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MeSH Terms
Blood Platelets*
Cell-Derived Microparticles / pathology*
Flow Cytometry / methods,  standards*
Observer Variation
Particle Size
Reference Standards

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine

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