Sequential CD34 cellfractionation by magnetophoresis in a magnetic dipole flow sorter

Abstract

Cell separation and fractionation based on fluorescent and magnetic labeling procedures are common tools in contemporary research. These techniques rely on binding of fluorophores or magnetic particles conjugated to antibodies to target cells. Cell surface marker expression levels within cell populations vary with progression through the cell cycle. In an earlier work we showed the reproducible magnetic fractionation (single pass) of the Jurkat cell line based on the population distribution of CD45 surface marker expression. Here we present a study on magnetic fractionation of a stem and progenitor cell (SPC) population using the established acute myelogenous leukemia cell line KG-1a as a cell model. The cells express a CD34 cell surface marker associated with the hematopoietic progenitor cell activity and the progenitor cell lineage commitment (related to the CD34 marker expression level). The CD34 expression level is approximately an order of magnitude lower than that of the CD45 marker, which required further improvements of the magnetic fractionation apparatus. The cells were immuno-magnetically labeled using a sandwich of anti CD34 antibody-phycoerythrin (PE) conjugate and anti PE magnetic nanobead and fractionated into eight components using a continuous flow dipole magnetophoresis apparatus. The CD34 marker expression distribution between sorted fractions was measured by quantitative PE flow cytometry (using QuantiBRITE™ PE calibration beads), and it was shown to be correlated with the cell magnetophoretic mobility distribution. A flow outlet addressing scheme based on the concept of the transport lamina thickness was used to control cell distribution between the eight outlet ports. The fractional cell distributions showed good agreement with numerical simulations of the fractionation based on the cell magnetophoretic mobility distribution in the unsorted sample.

Rare Cell Separation and Analysis by Magnetic Sorting

“…46 There is already a large body of data on similar systems such as field-flow fractionation, split-flow thin (SPLITT) channel fractionation and free-flow cell electrophoresis that is used for experimental design and data analysis, 47, 48 especially based on a useful concept of “transport lamina”. 12 Other potential future separation technologies using established physical principles include further refinement of filtration and dielectric fields. It is hoped that with the increasing interest in the need to obtain pure, or nearly pure cell subpopulations, especially when the targeted cells is rare, that cleaver new physical cell separation principles can be discovered to further push the field forward.…”

Fundamentals and application of magnetic particles in cell isolation and enrichment: a review

“…The same research group has illustrated that red blood cell removal, without magnetic tags, is possible using QMS (Moore et al, 2013, Moore et al, 2014, Jin et al, 2012, Zborowski et al, 2003). Using such a negative selection approach, hematopoietic stem cells have been enriched from blood (Jing et al, 2007, Tong et al, 2007, Schneider et al, 2010, Jin et al, 2012). Overall, the high throughput (> 10 6 cells/s) and high recovery capabilities (> 95%) of the QMS make it a strong magnetic separation technique (Zborowski and Chalmers, 2011, Moore et al, 1998, Schneider et al, 2006, Schneider et al, 2010).…”

Nanotechnology for the detection and kill of circulating tumor cells

“…It is an easy, rapid, and accurate method to separate RNA and tumor cells [51-53]. Herein, we introduce a novel CTC separation device applying the lateral magnetophoresis principle designed by Kim et al [23].…”