Flow cytometric detection of endothelial microparticles (EMP): Effects of centrifugation and storage alter with the phenotype studied

Ierssel, Sabrina H. van; Craenenbroeck, Emeline M. Van; Conraads, Viviane M.; Tendeloo, Viggo F. Van; Vrints, Christiaan J.; Jorens, Philippe G.; Hoymans, Vicky Y.

doi:10.1016/j.thromres.2009.12.019

Cited by 108 publications

(104 citation statements)

References 21 publications

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“…On fresh samples, a second centrifugation at 13,000g has been reported to decrease endothelial and annexin V1 microvesicles by about 50% in some studies (22,23), but not others (14,20,24). If plasma samples are going to be frozen, a single centrifugation step is inadequate to remove all platelets from the sample, double centrifugation is required to prevent 100% to 1,500% increases in platelet and annexin V1 microvesicle counts due to fragmentation of residual platelets in the sample (15,20,(25)(26)(27).…”

Section: Centrifugationmentioning

confidence: 96%

Measurement of microvesicle levels in human blood using flow cytometry

Chandler

2016

Cytometry Part B Clinical

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Microvesicles are fragments of cells released when the cells are activated, injured, or apoptotic. Analysis of microvesicle levels in blood has the potential to shed new light on the pathophysiology of many diseases. Flow cytometry is currently the only method that can simultaneously separate true lipid microvesicles from other microparticles in blood, determine the cell of origin and other microvesicle characteristics, and handle large numbers of clinical samples with a reasonable effort, but expanded use of flow cytometric measurement of microvesicle levels as a clinical and research tool requires improved, standardized assays. The goal of this review is to aid investigators in applying current best practices to microvesicle measurements. First pre-analytical factors are evaluated and data summarized for anticoagulant effects, sample transport and centrifugation. Next flow cytometer optimization is reviewed including interference from background in buffers and reagents, accurate microvesicle counting, swarm interference, and other types of coincidence errors, size calibration, and detection limits using light scattering, impedance and fluorescence. Finally current progress on method standardization is discussed and a summary of current best practices provided. V C 2016 Clinical Cytometry Society

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Section: Centrifugationmentioning

confidence: 96%

Measurement of microvesicle levels in human blood using flow cytometry

Chandler

2016

Cytometry Part B Clinical

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“…The time necessary for counting 2,500 events was determined and MP concentration (absolute number per microliter, n/μl) was determined from the formula: MP (n/μl)=(Number of events/ Volume sample analyzed)*(Total volume of the sample/Amount of platelet poor plasma), where the total volume of the ample was 500 μl, and 50 μl was the amount of platelet poor plasma. 21 The volume sample analyzed was calculated with the formula: 10/60*T, where T was the time of analysis expressed in seconds, 10 was the volume in μl analyzed by the cytometer in the unit of time (1 min), and 60 was the conversion factor from minutes to seconds.…”

Section: Blood Sampling and Mp Measurementmentioning

confidence: 99%

Differences in Microparticle Release in Patients With Acute Coronary Syndrome and Stable Angina

Biasucci

Porto

Vito

et al. 2012

Circ J

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Background: Microparticles (MP) are vesicles released from activated or apoptotic cells. Endothelial MP (EMP) are derived from injured endothelium, platelet MP (PMP) from activated platelets, and Annexin V positive MP (AMP) from apoptotic endothelial cells. The aim was to assess the release of MP and its association with inflammation and atherosclerotic burden. Methods and Results:AMP, EMP and PMP were measured on admission (Day 0) in 33 patients with stable angina (SA) and 43 patients with acute coronary syndrome (ACS) undergoing percutaneous coronary interventions (PCI). In SA, peripheral artery disease (PAD) was assessed by ultrasound examination. In 30 of the 76 patients (20 ACS and 10 SA), MP, high-sensitivity-C-reactive protein (hs-CRP), and troponin T (TnT) levels were also assessed 24 h (Day 1) and 48 h (Day 2) after PCI. AMP, EMP, and PMP were higher in ACS than in SA (all P<0.01). In the SA group, AMP, PMP, and EMP were similar in patients with or without PAD. In the ACS group, AMP increased until Day 2 (P=0.001), while EMP and PMP peaked on Day 1 (P<0.01) then decreased to baseline values. Day 2 AMP correlated with Day 2 TnT levels (r=0.43, P=0.01) while Day 1 EMP and PMP correlated with Day 1 hs-CRP (r=0.37, P=0.04 and r=0.33, P=0.05; respectively). Conclusions:Higher MP levels were observed in ACS than in SA. Atherosclerotic burden did not affect MP levels in stable patients. (Circ J 2012; 76: 2174 - 2182

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“…Endothelial micro particles (EMP) (12,13) as well as endothelial progenitor cells (EPC) (14) and circulating angiogenic cells (CAC) (15) emerged as markers of respectively endothelial damage and repair. All have been implicated in the process of childhood obesity-related endothelial dysfunction (16,17).…”

Section: Introductionmentioning

confidence: 99%