Microfluidic Separation of Cells against Multiple Markers

Abstract

Endothelial progenitor cells (EPCs) have the ability to repair damaged blood vessels and therefore are useful as precursors in the in vitro cultivation of vascular grafts. There is a need to sequester these cells from whole blood, an inherently heterogeneous mixture. The ability to isolate these rare cells from a complex blood suspension requires a separation process wherein the cells are isolated based on expression of multiple surface markers. Conventional methods for this separation require the attachment of antibody tags, typically fluorescent or magnetic, which may be undesirable for applications in tissue engineering. This study expands on previous work wherein a dissolvable hydrogel is functionalized with a capture ligand. Here this concept is expanded to a sequential enrichment of a target cell population against two surface markers specifically by using anti-CD34 to isolate the CD34+/ Flk1+ endothelial progenitor cells and subsequently using anti-Flk1 to deplete the CD34+/Flk1- hematopoietic stem cells. Such isolations can be accomplished by employing a microfluidic device for capture, two intermediate-stage devices designed to prepare the sample for further isolation, followed by a second capture stage. The strength of this method lies in the relative simplicity of the layout and the reasonable levels of cell recovery from a complex sample. Relevant applications include cell implantation-based therapeutics such as tissue engineering and molecular analysis.