BACKGROUND
Circulating endothelial cells (CECs) in the blood are rare but have been shown to be associated with various diseases. With the ratio of CECs to peripheral blood mononuclear cells (PBMCs) less than 1 part per thousand, their separation from PBMCs and detection are challenging. We present a means of detecting CECs from PBMCs via an economical microfluidic disk with a model cell system [human umbilical vein endothelial cells (HUVECs) in PBMCs], along with demonstration of its efficacy clinically.
METHODS
To enrich these rare cells, we used immunomagnetic beads and a tailor-made magnet on the disk. CEC-simulating HUVECs, as target cells, were stained with primary anti–CD146-phycoerythrin antibody and bound with secondary antibody on antiphycoerythrin magnetic beads. PBMCs served as nontarget cells and were labeled with anti–CD45-FITC antibody.
RESULTS
When hundreds of HUVECs were mixed in 106 PBMCs, 95% of spiked HUVECs were detected. This yield also held for 60 HUVEC in <104 PBMCs. We compared data from flow cytometry with that from the disk: CEC counts in 50 μL blood from patients with systemic lupus erythematosus were 61.1 (21.5), significantly higher (P < 0.01) than those of healthy donors, 31.2 (13.3).
CONCLUSIONS
The count of CECs is a suitable marker for symptoms of systemic lupus erythematosus. The microfluidic disk system should be a viable platform for detection of CECs.
Association of circulating endothelial cells with flow mediated vasodilation and disease activity in patients with systemic lupus erythematosus