Related Experiment Video
Updated: Aug 6, 2026
Isolation of Valvular Endothelial Cells
Published on: December 29, 2010
Isolation and enumeration of circulating endothelial cells by immunomagnetic isolation: proposal of a definition and
A Woywodt1, A D Blann, T Kirsch
1Department of Medicine, Division of Nephrology, Hannover Medical School, Hannover, Germany. woywodt.alexander@mh-hannover.de
Insights
This study establishes a consensus protocol for enumerating circulating endothelial cells (CECs), vital markers of vascular damage. The new method ensures reliable CEC identification and provides a working definition for these important cells.
Area of Science:
- Cardiovascular Research
- Cell Biology
- Biomarker Discovery
Background:
- Circulating endothelial cells (CECs) are key indicators of vascular damage in conditions like myocardial infarction and transplantation.
- Current CD146-driven immunomagnetic isolation methods lack standardized definitions and enumeration protocols for CECs.
Purpose of the Study:
- To optimize immunomagnetic isolation of CECs by evaluating key variables.
- To establish a clear definition for CECs.
- To propose a consensus protocol for CEC enumeration.
Main Methods:
- Developed a protocol using CD146-driven immunomagnetic isolation and Ulex Europaeus Lectin-1 staining.
- Conducted a multi-center evaluation of preanalytical (storage, anticoagulation) and analytical phases.
- Assessed the impact of density centrifugation on CEC recovery.
Main Results:
- Achieved unequivocal identification and acceptable reproducibility of CECs.
- Demonstrated that storage time affects cell numbers; recovery is reduced with citrate and density centrifugation.
- Provided technical recommendations and identified potential pitfalls.
- Formulated a working definition for CECs based on protocol experience and consensus.
Conclusions:
- This research is a significant advancement towards a consensus on CEC identification and enumeration.
- The proposed recommendations, based on multi-center experience, aim to guide future research and clinical applications.
Background:
Circulating endothelial cells (CECs) have been identified as markers of vascular damage in a variety of disorders, such as myocardial infarction, vasculitis, and transplantation. CD146-driven immunomagnetic isolation has gained widespread use, but the technique is hampered by the lack of a definition of CECs and the absence of a consensus for their enumeration.
Aim:
To evaluate several variables influencing immunomagnetic isolation of CECs, formulate a definition for CECs and propose a consensus protocol for their enumeration.
Methods:
We devised a protocol based on CD146-driven immunomagnetic isolation and a subsequent confirmatory step with Ulex-Europaeus-Lectin-1 staining. In a multi-center effort, we evaluated the preanalytical and analytical phases of this protocol. We evaluated the effects of storage, anticoagulation and density centrifugation, and compiled previous experience with this technique.
Results:
Our protocol permitted unequivocal identification of CECs with acceptable reproducibility. There was an effect of storage time in that median cell numbers declined to only 87.5% of their baseline values during 24 h of storage at 4 degrees C. Recovery was lower with citrate than with ethylene-diamine tetra-acetic acid after 4 h of storage; density centrifugation was also associated with lower recovery. We provide a comprehensive list of technical recommendations and potential pitfalls. Finally, based on our experience with this protocol and a recent consensus workshop, we formulated a working definition for CECs.
Conclusion:
Our work represents an important step toward consensus regarding the CECs. Our recommendations represent the experience of three major centers and should now be scrutinized by others in the field.
