| Quantification of circulating mature endothelial cells using a whole blood four-color flow cytometric assay. | |
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MedLine Citation:
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PMID: 18662694 Owner: NLM Status: MEDLINE |
Abstract/OtherAbstract:
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Circulating endothelial cells (CEC) are currently proposed as a potential biomarker for measuring the impact of anti-angiogenic treatments in cancer. However, the lack of consensus on the appropriate method of CEC measurement has led to conflicting data in cancer patients. A validated assay adapted for evaluating the clinical utility of CEC in large cohorts of patients undergoing anti-angiogenic treatments is needed. We developed a four-color flow cytometric assay to measure CEC as CD31(+), CD146(+), CD45(-), 7-amino-actinomycin-D (7AAD)(-) events in whole blood. The distinctive features of the assay are: (1) staining of 1 ml whole blood, (2) use of a whole blood IgPE control to measure accurately background noise, (3) accumulation of a large number of events (almost 5 10(6)) to ensure statistical analysis, and (4) use of 10 microm fluorescent microbeads to evaluate the event size. Assay reproducibility was determined in duplicate aliquots of samples drawn from 20 metastatic cancer patients. Assay linearity was tested by spiking whole blood with low numbers of HUVEC. Five-color flow cytometric experiments with CD144 were performed to confirm the endothelial origin of the cells. CEC were measured in 20 healthy individuals and 125 patients with metastatic cancer. Reproducibility was good between duplicate aliquots (r(2)=0.948, mean difference between duplicates of 0.86 CEC/ml). Detected HUVEC correlated with spiked HUVEC (r(2)=0.916, mean recovery of 100.3%). Co-staining of CD31, CD146 and CD144 confirmed the endothelial nature of cells identified as CEC. Median CEC levels were 6.5/ml (range, 0-15) in healthy individuals and 15.0/ml (range, 0-179) in patients with metastatic carcinoma (p<0.001). The assay proposed here allows reproducible and sensitive measurement of CEC by flow cytometry and could help evaluate CEC as biomarkers of anti-angiogenic therapies in large cohorts of patients. |
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Authors:
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Nathalie Jacques; Nadege Vimond; Rosa Conforti; Franck Griscelli; Yann Lecluse; Agnes Laplanche; David Malka; Philippe Vielh; Françoise Farace |
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Publication Detail:
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Type: Journal Article Date: 2008-07-26 |
Journal Detail:
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Title: Journal of immunological methods Volume: 337 ISSN: 0022-1759 ISO Abbreviation: J. Immunol. Methods Publication Date: 2008 Sep |
Date Detail:
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Created Date: 2008-08-25 Completed Date: 2008-10-07 Revised Date: - |
Medline Journal Info:
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Nlm Unique ID: 1305440 Medline TA: J Immunol Methods Country: Netherlands |
Other Details:
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Languages: eng Pagination: 132-43 Citation Subset: IM |
Affiliation:
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Translational Research Laboratory, Gustave Roussy Institute, 94805 Villejuif, France. |
Export Citation:
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| MeSH Terms | |
Descriptor/Qualifier:
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Angiogenesis Inhibitors
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therapeutic use* Antigens, CD / analysis Biological Markers Blood Cell Count / methods* Cadherins / analysis Endothelial Cells / cytology* Flow Cytometry / methods* Humans Neoplasm Metastasis Neoplasms / blood Reproducibility of Results Staining and Labeling |
| Chemical | |
Reg. No./Substance:
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0/Angiogenesis Inhibitors; 0/Antigens, CD; 0/Biological Markers; 0/Cadherins; 0/cadherin 5 |
From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine
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