Cell tracking velocimetry as a tool for defining saturation binding of magnetically conjugated antibodies

Abstract

Background: Continuous flow immunomagnetic separation is an attractive alternative to current batch mode immunomagnetic separation methods because it is capable of high sorting speeds at mild cell conditions, and grants the operator better control of separation process. The control of the separation is dependent on knowledge of the amount of magnetic label attached to the cell (magnetic labeling intensity), however. Determination of the magnetic labeling is accomplished by measuring cell magnetophoretic mobility using a newly developed technique of Cell Tracking Velocimetry (CTV). Methods: Flow cytometry was used to define the antibody binding characteristics of a fluorescently tagged primary antibody. Subsequently, CTV was used to measure antibody‐binding characteristics of a magnetically tagged secondary antibody. Results: The results of this study show that CTV is capable of providing valuable information concerning the cell labeling by magnetically tagged antibodies. It was demonstrated that the magnetically conjugated antibody binding curve exhibits the same exponential increase to saturation characteristics as that seen with the fluorescently tagged antibody. Further, it was shown that the intensity of the secondary magnetic labeling is directly proportional to the intensity of the primary fluorescent label. Conclusions: CTV is an accurate tool for evaluation of magnetically conjugated antibodies. The ability to determine the intensity of magnetic labeling is necessary for the development of continuous flow immunomagnetic separations based on cell magnetophoresis. © 2005 Wiley‐Liss, Inc.

Sequential CD34 cellfractionation by magnetophoresis in a magnetic dipole flow sorter

“…This is consistent with the double label binding mechanism for which the presence of a PE moiety on a cell is necessary for magnetic particle binding. Increase in PE molecules bound per cell is therefore correlated with increase in cell magnetization, and hence, magnetophoretic mobility 44. Furthermore, the mean number of PE molecules in the initial cell sample fell between the upper and lower limits of the sorted fractions, as expected (Figure 3a).…”

Differences in magnetically induced motion of diamagnetic, paramagnetic, and superparamagnetic microparticles detected by cell tracking velocimetry

“…A cell labeled with magnetic nanoparticles acquires a MM that is quantitatively related to the physio-chemical properties of the cell-label complex and the suspending fluid, such as the magnetic susceptibility of the nanoparticles, the hydrodynamic diameter of the cell-label complex, the antibody binding capacity of the cell, and the viscosity of the fluid medium 11. The MM distribution in the cell population depends on the selectivity and specificity of the targeting antibodies and the cell surface antigen expression 12,13…”

The Fabrication and Application Mechanism of Microfluidic Systems for High Throughput Biomedical Screening: A Review

“…However, the intrinsic properties of the cells are sometimes not enough to be separated by magnetophoresis. The cells could be labeled by special antibody-magnetic particle conjugates to increase the magnetic susceptibility [94,[97][98][99][100][101]. Schneider et al labeled cells with antibodies conjugated to magnetic nanospheres.…”