A current roadblock for human stem cell therapies is the lack of efficient cell sorting methodologies to produce purified cells for transplantation. Stem cell transplants should contain purified cells designed to provide functional recovery but lack cells that can form tumors, making pure cell populations a necessity for the stem cell field. Conventional techniques rely on the presence of specific cell surface markers, which have not been identified for many stem cells of potential therapeutic interest. We have addressed this problem by developing a new sorting method using microfluidics and a technology termed “dielectrophoresis” (DEP) that utilizes electric fields to separate cells. Our data show that DEP distinguishes neural stem/progenitor cells (NSPCs) from further differentiated cells, can be used to enrich NSPCs biased to make neurons, and has no deleterious effects on human NSPCs.
DEP is likely to be an optimal method for sorting stem cells prior to transplantation for several reasons. DEP-based cell sorting requires no labeling, so cells isolated in this manner will not be coated by antibodies immediately prior to transplantation, potentially reducing immune-mediated rejection of sorted cells. Furthermore, cells can be sorted by DEP through multiple channels with distinct DEP force profiles to refine separation in a single sort; a similar strategy using other methods would require additional time to label the cells with antibodies against a different set of markers. Our data show that DEP does not alter or harm human NSPCs and the recovery and survival of these cells after DEP is superior to that of cells isolated by other methods. Importantly, initial studies show that isolation of NSPCs by DEP generates greater enrichment of certain cell types than other methods.
Current sorting methodologies using cell surface markers are insufficient for purifying many stem cell populations, including neuron-restricted progenitors in the neural lineage, due to a lack of necessary specificity of available markers. The goal of our proposal is to improve the sorting of stem cells prior to transplantation by developing a novel and label-free method complementary to currently used marker-based techniques.
The goal of this project is to determine whether a novel strategy using dielectrophoresis (DEP) can serve as a complementary and alternative approach to marker-based separation of stem cell subpopulations to be used for transplantation. In the course of these studies, we expect to remove tumor-promoting cells from neural human progenitor cells to improve their safety for transplantation. We also hope to make purified populations of cells that can replace specific central nervous system cells. We hope this label-free method for isolating stem cell subpopulations will greatly increase the speed of stem cell therapeutics in California.