AustralieFrV1, PubMed, Checkpoint, bibRecord, 002A79

Immuno-analysis of microparticles: probing at the limits of detection.

Identifieur interne : 002A79 ( PubMed/Checkpoint ); précédent : 002A78; suivant : 002A80

Immuno-analysis of microparticles: probing at the limits of detection.

Auteurs : Sharissa L. Latham [Australie] ; Natalia Tiberti [Australie] ; Naveena Gokoolparsadh [Australie] ; Karen Holdaway [Australie] ; Pierre Olivier Couraud [France] ; Georges E R. Grau [Australie] ; Valery Combes [Australie]

Source :

Scientific reports [ 2045-2322 ] ; 2015.

RBID : pubmed:26553743

Descripteurs français

English descriptors

KwdEn :
- Annexin A5 (immunology), Annexin A5 (metabolism), Antigens, CD (immunology), Antigens, CD (metabolism), Antigens, Surface (immunology), Antigens, Surface (metabolism), Cell Line, Cell-Derived Microparticles (chemistry), Cell-Derived Microparticles (immunology), Cell-Derived Microparticles (metabolism), Endoglin, Flow Cytometry, Gold (chemistry), Humans, Intercellular Adhesion Molecule-1 (immunology), Intercellular Adhesion Molecule-1 (metabolism), Limit of Detection, Microscopy, Immunoelectron, Milk Proteins (immunology), Milk Proteins (metabolism), Phosphatidylserines (immunology), Phosphatidylserines (metabolism), Receptors, Cell Surface (immunology), Receptors, Cell Surface (metabolism).
MESH :
- chemical , chemistry : Gold.
- chemical , immunology : Annexin A5, Antigens, CD, Antigens, Surface, Intercellular Adhesion Molecule-1, Milk Proteins, Phosphatidylserines, Receptors, Cell Surface.
- chemical , metabolism : Annexin A5, Antigens, CD, Antigens, Surface, Intercellular Adhesion Molecule-1, Milk Proteins, Phosphatidylserines, Receptors, Cell Surface.
- chemistry : Cell-Derived Microparticles.
- immunology : Cell-Derived Microparticles.
- metabolism : Cell-Derived Microparticles.
- Cell Line, Endoglin, Flow Cytometry, Humans, Limit of Detection, Microscopy, Immunoelectron.

Abstract

Microparticle (MP) research is clouded by debate regarding the accuracy and validity of flow cytometry (FCM) as an analytical methodology, as it is influenced by many variables including the pre-analytical conditions, instruments physical capabilities and detection parameters. This study utilises a simplistic in vitro system for generating MP, and through comparative analysis with immuno-electron microscopy (Immuno-EM) assesses the strengths and limitations of probe selection and high-sensitivity FCM. Of the markers examined, MP were most specifically labelled with phosphatidylserine ligands, annexin V and lactadherin, although only ~60% MP are PS positive. Whilst these two ligands detect comparable absolute MP numbers, they interact with the same population in distinct manners; annexin V binding is enhanced on TNF induced MP. CD105 and CD54 expression were, as expected, consistent and enhanced following TNF activation respectively. Their labelling however accounted for as few as 30-40% of MP. The greatest discrepancies between FCM and I-EM were observed in the population solely labelled for the surface antigen. These findings demonstrate that despite significant improvements in resolution, high-sensitivity FCM remains limited in detecting small-size MP expressing low antigen levels. This study highlights factors to consider when selecting endothelial MP probes, as well as interpreting and representing data.

DOI: 10.1038/srep16314
PubMed: 26553743

Affiliations:

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Le document en format XML

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<front><div type="abstract" xml:lang="en">Microparticle (MP) research is clouded by debate regarding the accuracy and validity of flow cytometry (FCM) as an analytical methodology, as it is influenced by many variables including the pre-analytical conditions, instruments physical capabilities and detection parameters. This study utilises a simplistic in vitro system for generating MP, and through comparative analysis with immuno-electron microscopy (Immuno-EM) assesses the strengths and limitations of probe selection and high-sensitivity FCM. Of the markers examined, MP were most specifically labelled with phosphatidylserine ligands, annexin V and lactadherin, although only ~60% MP are PS positive. Whilst these two ligands detect comparable absolute MP numbers, they interact with the same population in distinct manners; annexin V binding is enhanced on TNF induced MP. CD105 and CD54 expression were, as expected, consistent and enhanced following TNF activation respectively. Their labelling however accounted for as few as 30-40% of MP. The greatest discrepancies between FCM and I-EM were observed in the population solely labelled for the surface antigen. These findings demonstrate that despite significant improvements in resolution, high-sensitivity FCM remains limited in detecting small-size MP expressing low antigen levels. This study highlights factors to consider when selecting endothelial MP probes, as well as interpreting and representing data.</div>
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<Abstract><AbstractText>Microparticle (MP) research is clouded by debate regarding the accuracy and validity of flow cytometry (FCM) as an analytical methodology, as it is influenced by many variables including the pre-analytical conditions, instruments physical capabilities and detection parameters. This study utilises a simplistic in vitro system for generating MP, and through comparative analysis with immuno-electron microscopy (Immuno-EM) assesses the strengths and limitations of probe selection and high-sensitivity FCM. Of the markers examined, MP were most specifically labelled with phosphatidylserine ligands, annexin V and lactadherin, although only ~60% MP are PS positive. Whilst these two ligands detect comparable absolute MP numbers, they interact with the same population in distinct manners; annexin V binding is enhanced on TNF induced MP. CD105 and CD54 expression were, as expected, consistent and enhanced following TNF activation respectively. Their labelling however accounted for as few as 30-40% of MP. The greatest discrepancies between FCM and I-EM were observed in the population solely labelled for the surface antigen. These findings demonstrate that despite significant improvements in resolution, high-sensitivity FCM remains limited in detecting small-size MP expressing low antigen levels. This study highlights factors to consider when selecting endothelial MP probes, as well as interpreting and representing data.</AbstractText>
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<Initials>SL</Initials>
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</AffiliationInfo>
<AffiliationInfo><Affiliation>Institute for Biophysical Chemistry, Hannover Medical School, Hannover, Germany.</Affiliation>
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</AffiliationInfo>
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<Initials>V</Initials>
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</AffiliationInfo>
<AffiliationInfo><Affiliation>Faculty of Science, School of Life Sciences, University of Technology Sydney, NSW, Australia.</Affiliation>
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<orgName><li>Université de Sydney</li>
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<tree><country name="Australie"><region name="Nouvelle-Galles du Sud"><name sortKey="Latham, Sharissa L" sort="Latham, Sharissa L" uniqKey="Latham S" first="Sharissa L" last="Latham">Sharissa L. Latham</name>
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<name sortKey="Combes, Valery" sort="Combes, Valery" uniqKey="Combes V" first="Valery" last="Combes">Valery Combes</name>
<name sortKey="Gokoolparsadh, Naveena" sort="Gokoolparsadh, Naveena" uniqKey="Gokoolparsadh N" first="Naveena" last="Gokoolparsadh">Naveena Gokoolparsadh</name>
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<name sortKey="Tiberti, Natalia" sort="Tiberti, Natalia" uniqKey="Tiberti N" first="Natalia" last="Tiberti">Natalia Tiberti</name>
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<country name="France"><region name="Île-de-France"><name sortKey="Couraud, Pierre Olivier" sort="Couraud, Pierre Olivier" uniqKey="Couraud P" first="Pierre Olivier" last="Couraud">Pierre Olivier Couraud</name>
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	Serveur d'exploration sur les relations entre la France et l'Australie
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Immuno-analysis of microparticles: probing at the limits of detection.

Immuno-analysis of microparticles: probing at the limits of detection.

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