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Impact of sample processing on the measurement of circulating microparticles: storage and centrifugation parameters

  • Virtudes Vila-Liante , Verónica Sánchez-López , Vicenta Martínez-Sales , Luis A. Ramón-Nuñez , Elena Arellano-Orden , Alejandra Cano-Ruiz , Francisco J. Rodríguez-Martorell , Lin Gao and Remedios Otero-Candelera EMAIL logo
Published/Copyright: May 6, 2016
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Clinical Chemistry and Laboratory Medicine (CCLM)
From the journal Clinical Chemistry and Laboratory Medicine (CCLM) Volume 54 Issue 11

Abstract

Background:

Microparticles (MPs) have been shown to be markers of cellular activation and interactions. Pre-analytical conditions such as the centrifugation protocol and sample storage conditions represent an important source of variability in determining MPs values. The objectives of this study were to evaluate the influence of sample storage conditions and centrifugation speed and temperature on the determination of MPs in plasma.

Methods:

Citrate-anticoagulated blood samples obtained from 21 healthy subjects were centrifuged under four different protocols involving different speeds (2500 g or 1500 g) and temperatures (4 °C or 20 °C) to isolate platelet-poor plasma (PPP). The number of MPs in fresh and frozen-thawed PPP were analyzed by flow cytometry, and MPs-mediated procoagulant activity was determined by a thrombin generation test and phospholipid-dependent procoagulant tests.

Results:

The number of MPs and their procoagulant activity were affected by freeze-thaw cycling and centrifugation speed but not by centrifugation temperature. Sample freezing increased MPs number (six-fold) and thrombin generation (four-fold), and decreased clotting time (two-fold). Low centrifugation speed caused an increase in MPs number and a parallel increase in MP-mediated procoagulant activity.

Conclusions:

Sample storage conditions and centrifugation speed are important processing conditions affecting MPs number and activity. Before any study, the protocol for MPs isolation should be optimized to ensure a reliable characterization of MPs, which could provide important information for diagnostic purposes and for understanding the pathogenesis of diseases.

Keywords: cell derived microparticles; centrifugation; flow cytometry; freezing; pre-analytics; standardization
Corresponding author: Remedios Otero-Candelera, MD, PhD, Instituto de Biomedicina de Sevilla (IBiS). Hospital Universitario Virgen del Rocio/CSIC/Universidad de Sevilla, 41013 Seville, Spain; Centro de Investigación Biomédica en Red de Enfermedades Respiratorias (CIBERES), Spain; and Avda Manuel Siurot s/n, 41013 Seville, Spain, Phone: +34 955013159, Fax: +34 955013166
aVirtudes Vila-Liante and Verónica Sánchez-López contributed equally to this work and both should be considered first author.

Funding source: Instituto de Salud Carlos III

Award Identifier / Grant number: PI11/02308

Funding statement: This work was supported by research grants from the FEDER fundings and Plan Nacional de Investigación Científica, Desarrollo e Innovación Tecnológica [Instituto de Salud Carlos III, Fondo de Investigación Sanitaria (PI11/02308)], Red RIC (RD12/0042/0029), Junta de Andalucia (CVI-6654) and by the Fundación Investigación Hospital La Fe, Spain.

  1. Author contributions: All the authors have accepted responsibility for the entire content of this submitted manuscript and approved submission.

  2. Research funding: This work was supported by research grants from the FEDER fundings and Plan Nacional de Investigación Científica, Desarrollo e Innovación Tecnológica [Instituto de Salud Carlos III, Fondo de Investigación Sanitaria (PI11/02308)], Red RIC (RD12/0042/0029), Junta de Andalucia (CVI-6654) and by the Fundación Investigación Hospital La Fe, Spain.

  3. Employment or leadership: None declared.

  4. Honorarium: None declared.

  5. Competing interests: The funding organization(s) played no role in the study design; in the collection, analysis, and interpretation of data; in the writing of the report; or in the decision to submit the report for publication.

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Received: 2016-1-15
Accepted: 2016-3-30
Published Online: 2016-5-6
Published in Print: 2016-11-1

©2016 Walter de Gruyter GmbH, Berlin/Boston

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https://doi.org/10.1515/cclm-2016-0036
Keywords for this article
cell derived microparticles; centrifugation; flow cytometry; freezing; pre-analytics; standardization

Articles in the same Issue

  1. Frontmatter
  2. Editorials
  3. CCLM Award for the Most Cited Paper Recently Published
  4. Laboratory economics. Risk or opportunity?
  5. Reviews
  6. Molecular diagnosis and precision medicine in allergy management
  7. The insulin autoimmune syndrome (IAS) as a cause of hypoglycaemia: an update on the pathophysiology, biochemical investigations and diagnosis
  8. Opinion Paper
  9. Role of microsatellite instability, immunohistochemistry and mismatch repair germline aberrations in immunosuppressed transplant patients: a phenocopy dilemma in Muir-Torre syndrome
  10. Genetics and Molecular Diagnostics
  11. Time and tumor type (primary or metastatic) do not influence the detection of BRAF/NRAS mutations in formalin fixed paraffin embedded samples from melanomas
  12. False low holotranscobalamin levels in a patient with a novel TCN2 mutation
  13. General Clinical Chemistry and Laboratory Medicine
  14. Quality performance of laboratory testing in pharmacies: a collaborative evaluation
  15. Cost evaluation of clinical laboratory in Taiwan’s National Health System by using activity-based costing
  16. Impact of sample processing on the measurement of circulating microparticles: storage and centrifugation parameters
  17. HbA1c: EQA in Germany, Belgium and the Netherlands using fresh whole blood samples with target values assigned with the IFCC reference system
  18. Iohexol clearance in unstable critically ill patients: a tool to assess glomerular filtration rate
  19. Characteristics of Chinese patients with antiphospholipid syndrome and the ability of lupus anticoagulant assays to identify them
  20. Reference Values and Biological Variations
  21. Distribution of soluble suppression of tumorigenicity 2 (sST2), N-terminal pro-brain natriuretic peptide (NT-proBNP), high sensitive troponin I and high-sensitive troponin T in umbilical cord blood
  22. Hematology and Coagulation
  23. Evaluation of Mindray BC-6800 body fluid mode for automated cerebrospinal fluid cell counting
  24. Cancer Diagnostics
  25. Increased sialylation and reduced fucosylation of exfoliated cervical cells are potential markers of carcinogenesis in the cervix
  26. Cardiovascular Diseases
  27. High incidence of macrotroponin I with a high-sensitivity troponin I assay
  28. Infectious Diseases
  29. Prospective evaluation of biomarkers for prediction of quality of life in community-acquired pneumonia
  30. Letter to the Editor
  31. The use of extra-analytical phase quality indicators by clinical laboratories: the results of an international survey
  32. Quality of reporting of diagnostic test accuracy studies in medical laboratory journals
  33. Impact of under-filled blood collection tubes containing K2EDTA and K3EDTA as anticoagulants on automated complete blood count (CBC) testing
  34. Stability of plasma albumin depends on measurement method
  35. Evaluation of screening method for Bence Jones protein analysis
  36. Reference intervals for the Kryptor second-generation chromogranin A assay
  37. Evaluation of an automated urinary iodine measurement using AU5800 analyzer with AutoLab Iodine reagent
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  39. The use of a “gray zone” considering measurement uncertainty in pharmacological tests. The serum growth hormone stimulation test as an example
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