A flow cytometric method for characterization of circulating cell-derived microparticles in plasma

Morten Hjuler Nielsen, Henning Beck-Nielsen, Morten Nørgaard Andersen, Aase Handberg

Research output: Contribution to journalJournal articleResearchpeer-review

Abstract

BACKGROUND AND AIM: Previous studies on circulating microparticles (MPs) indicate that the majority of MPs are of a size below the detection limit of most standard flow cytometers. The objective of the present study was to establish a method to analyze MP subpopulations above the threshold of detection of a new generation BD FACSAria™ III digital flow cytometer.

METHODS: We analyzed MP subpopulations in plasma from 24 healthy individuals (9 males and 15 females). MPs were identified according to their size (<1.0-µm), by Lactadherin-FITC labelling, and by exposure of cell-specific markers. The sensitivity of the flow cytometer was tested against that of a previous-generation instrument FC500. Reproducibility of the FACSAria and our set-up was investigated, and the percentage of phosphatidylserine (PS) exposing MPs binding Lactadherin was determined.

RESULTS: By using a flow cytometric approach we identified and quantitated MPs derived from platelets, monocytes, erythrocytes and endothelial cells. In addition, levels of tissue factor-positive MPs were determined. The FACSAria demonstrated improved sensitivity and increased MP detection range compared to the FC500 instrument. The reproducibility of PS+PMP and PS+MP measurements was 11.7 and 23.2%, respectively. When expressed as a percentage of total MPs, the PS-positive MP population represented 15.1±5.5%, and PS-positive MPs were significantly increased in men.

CONCLUSION: We have established a method to measure MPs above the detection limit of a new generation flow cytometer and derived from a number of cell-types in a healthy population of men and women.

Original languageEnglish
JournalJournal of Extracellular Vesicles
Volume3
ISSN2001-3078
DOIs
Publication statusPublished - 2014

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Cell-Derived Microparticles
Phosphatidylserines
Limit of Detection
Fluorescein-5-isothiocyanate
Population

Cite this

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title = "A flow cytometric method for characterization of circulating cell-derived microparticles in plasma",
abstract = "BACKGROUND AND AIM: Previous studies on circulating microparticles (MPs) indicate that the majority of MPs are of a size below the detection limit of most standard flow cytometers. The objective of the present study was to establish a method to analyze MP subpopulations above the threshold of detection of a new generation BD FACSAria™ III digital flow cytometer.METHODS: We analyzed MP subpopulations in plasma from 24 healthy individuals (9 males and 15 females). MPs were identified according to their size (<1.0-µm), by Lactadherin-FITC labelling, and by exposure of cell-specific markers. The sensitivity of the flow cytometer was tested against that of a previous-generation instrument FC500. Reproducibility of the FACSAria and our set-up was investigated, and the percentage of phosphatidylserine (PS) exposing MPs binding Lactadherin was determined.RESULTS: By using a flow cytometric approach we identified and quantitated MPs derived from platelets, monocytes, erythrocytes and endothelial cells. In addition, levels of tissue factor-positive MPs were determined. The FACSAria demonstrated improved sensitivity and increased MP detection range compared to the FC500 instrument. The reproducibility of PS+PMP and PS+MP measurements was 11.7 and 23.2{\%}, respectively. When expressed as a percentage of total MPs, the PS-positive MP population represented 15.1±5.5{\%}, and PS-positive MPs were significantly increased in men.CONCLUSION: We have established a method to measure MPs above the detection limit of a new generation flow cytometer and derived from a number of cell-types in a healthy population of men and women.",
author = "Nielsen, {Morten Hjuler} and Henning Beck-Nielsen and Andersen, {Morten N{\o}rgaard} and Aase Handberg",
year = "2014",
doi = "10.3402/jev.v3.20795",
language = "English",
volume = "3",
journal = "Journal of Extracellular Vesicles",
issn = "2001-3078",
publisher = "Co-Action Publishing",

}

A flow cytometric method for characterization of circulating cell-derived microparticles in plasma. / Nielsen, Morten Hjuler; Beck-Nielsen, Henning; Andersen, Morten Nørgaard; Handberg, Aase.

In: Journal of Extracellular Vesicles, Vol. 3, 2014.

Research output: Contribution to journalJournal articleResearchpeer-review

TY - JOUR

T1 - A flow cytometric method for characterization of circulating cell-derived microparticles in plasma

AU - Nielsen, Morten Hjuler

AU - Beck-Nielsen, Henning

AU - Andersen, Morten Nørgaard

AU - Handberg, Aase

PY - 2014

Y1 - 2014

N2 - BACKGROUND AND AIM: Previous studies on circulating microparticles (MPs) indicate that the majority of MPs are of a size below the detection limit of most standard flow cytometers. The objective of the present study was to establish a method to analyze MP subpopulations above the threshold of detection of a new generation BD FACSAria™ III digital flow cytometer.METHODS: We analyzed MP subpopulations in plasma from 24 healthy individuals (9 males and 15 females). MPs were identified according to their size (<1.0-µm), by Lactadherin-FITC labelling, and by exposure of cell-specific markers. The sensitivity of the flow cytometer was tested against that of a previous-generation instrument FC500. Reproducibility of the FACSAria and our set-up was investigated, and the percentage of phosphatidylserine (PS) exposing MPs binding Lactadherin was determined.RESULTS: By using a flow cytometric approach we identified and quantitated MPs derived from platelets, monocytes, erythrocytes and endothelial cells. In addition, levels of tissue factor-positive MPs were determined. The FACSAria demonstrated improved sensitivity and increased MP detection range compared to the FC500 instrument. The reproducibility of PS+PMP and PS+MP measurements was 11.7 and 23.2%, respectively. When expressed as a percentage of total MPs, the PS-positive MP population represented 15.1±5.5%, and PS-positive MPs were significantly increased in men.CONCLUSION: We have established a method to measure MPs above the detection limit of a new generation flow cytometer and derived from a number of cell-types in a healthy population of men and women.

AB - BACKGROUND AND AIM: Previous studies on circulating microparticles (MPs) indicate that the majority of MPs are of a size below the detection limit of most standard flow cytometers. The objective of the present study was to establish a method to analyze MP subpopulations above the threshold of detection of a new generation BD FACSAria™ III digital flow cytometer.METHODS: We analyzed MP subpopulations in plasma from 24 healthy individuals (9 males and 15 females). MPs were identified according to their size (<1.0-µm), by Lactadherin-FITC labelling, and by exposure of cell-specific markers. The sensitivity of the flow cytometer was tested against that of a previous-generation instrument FC500. Reproducibility of the FACSAria and our set-up was investigated, and the percentage of phosphatidylserine (PS) exposing MPs binding Lactadherin was determined.RESULTS: By using a flow cytometric approach we identified and quantitated MPs derived from platelets, monocytes, erythrocytes and endothelial cells. In addition, levels of tissue factor-positive MPs were determined. The FACSAria demonstrated improved sensitivity and increased MP detection range compared to the FC500 instrument. The reproducibility of PS+PMP and PS+MP measurements was 11.7 and 23.2%, respectively. When expressed as a percentage of total MPs, the PS-positive MP population represented 15.1±5.5%, and PS-positive MPs were significantly increased in men.CONCLUSION: We have established a method to measure MPs above the detection limit of a new generation flow cytometer and derived from a number of cell-types in a healthy population of men and women.

U2 - 10.3402/jev.v3.20795

DO - 10.3402/jev.v3.20795

M3 - Journal article

VL - 3

JO - Journal of Extracellular Vesicles

JF - Journal of Extracellular Vesicles

SN - 2001-3078

ER -