Magnetic Beads (MBs) cell sorting is a highly specific cell sorting technology that integrates fields of immunology, cell biology, and magnetism. Its high specificity comes from the specific recognition between antigens and antibodies. From laboratory to clinical, from small-scale to large-scale, from common cells to rare cells and complex cell subgroups, as well as from human and mouse cells to cells of other germ lines, MBs cell sorting technology has become a standard method for high quality cell sorting. In addition, because of its excellent specificity and separation efficiency, MBs cell sorting is widely used for various biomedical applications including cell biology, disease diagnostics and drug development.
Mechanism of Magnetic Beads (MBs) Cell Sorting
The MBs that have been labelled with the primary antibody specifically bind to the corresponding molecules on the cell surface, or the MBs that have been labelled with the secondary antibody bind to the primary antibody that has been previously bound to the cell surface molecules. Then the MBs carry the cells that bound to them and are adsorbed under the separation column/test tube to realize the separation.
MBs cell sorting can separate very pure cells from complex mixtures within minutes. When using nanoscale MBs for cell sorting, the size and composition will make it biodegradable without activating the cells or affecting the vitality of cells, and the physiological functions of cells are also unchanged. Magnetically labeled cells can be immediately used for analysis and subsequent experiments. This process can be divided into two types according to the different binding ways of MBs: positive sorting and negative sorting.
Methods of Magnetic Beads (MBs) Cell Sorting
Positive Sorting
Positive sorting uses antibodies or ligands to directly label targeted cells for sorting. The targeted cells are marked by antibodies and bound to MBs, and then placed under the external magnetic field. Due to the marked MBs, the targeted cells can be absorbed under the magnetic field to realize separation. The cells separated by positive sorting can be used immediately for analysis and subsequent experiments.
Advantages of positive sorting:
- Fast speed andsimple operation.
- High purity and high recovery rate.
- Separated cells can be used immediately for subsequent analysis or experiments
Negative Sorting
Negative sorting refers to the usage of antibody coupled MBs to label non-targeted cells. Because of the magnetism, non-targeted cells will be absorbed by the external magnetic field while the targeted cells are remained, thus realizing separation.
Advantages of negative sorting application:
- Remove undesired cells.
- For those cells that lack of specific antibodies, such as tumor cells.
- The antibody only targets all non-targeted cellsand does not stimulate the targeted cells.
- Negative sorting is always a part of customized sorting.
Modes of Magnetic Beads (MBs) Cell Sorting
- Column Cell Sorting
The column cell sorting means that cells labeled with MBs flow through a separation column placed in a magnetic field to separate the target cells. The separation column is made up of tiny iron particles. When these iron particles are placed in a magnetic field, they will generate a local magnetic field that can adsorb cells labelled with MBs. When using positive sorting, non-targeted cells not labeled with MBs can pass through the column, while targeted cells labeled with MBs are remained. The targeted cells can be eluted with buffer and collected after removing the column from the magnetic field.
- Column-free Cell Sorting
The column-free cell sorting is a process without columns. It refers to directly placing the test tube filled with cell populations containing targeted cells and non-targeted cells into a magnetic field. The targeted cells labelled with MBs are adsorbed under the exposure of the magnetic field. By contrast, those unlabeled cells will be removed by pouring or pipetting. After removing the test tube from the magnetic field, the targeted cells can be suspended in the test tube and collected.
Advantages of Magnetic Beads (MBs) Cell Sorting
MBs cell sorting can separate very pure cells from complex mixtures within minutes. When using nanoscale MBs for cell sorting, its size and composition will make it biodegradable without activating the cells or affecting the vitality of cells, and the physiological functions of cells are also unchanged. Magnetically labeled cells can be immediately used for analysis and subsequent experiments. MBs cell sorting is one of the commonly used methods for researchers to isolate specific cell populations due to the following advantages:
Precautions for Magnetic Beads (MBs) Cell Sorting
- If there are adherent cells in the cell populations that need to be sorted, it is recommended to culture and remove them before sorting.
- Dead cells need to be removed before sorting because antibodies labelled with MBs often have non-specific binding interaction to them.
- Before loading into the separation column, fully vibrating to suspend the cells and break up cell clumps is recommended.
- When using the column cell sorting, it is recommended to apply vacuum to filter the water to reduce bubbles in the water and prevent the separation column from being blocked by bubbles.
- After extending the dropper to the bottom wall of the tube, the cell suspension can be added to the separation column to avoid low purity. When washing the column, add the washing liquid after the previous liquid is completely drained.
- The amounts of sorted cells should not be excessive.
- Incubation time and temperature should be controlled. Extending the incubation time and increasing the temperature will increase non-specific binding.
References
1. Holt, L. M.; Olsen, M. L., Novel Applications of Magnetic Cell Sorting to Analyze Cell-Type Specific Gene and Protein Expression in the Central Nervous System. PLOS ONE 2016, 11 (2), e0150290.
2. Philippova, O.; Barabanova, A.; et al. Magnetic polymer beads: Recent trends and developments in synthetic design and applications. European polymer journal 2011, 47 (4), 542-559.
3. Pankhurst, Q. A.; Connolly, J.;et al. Applications of magnetic nanoparticles in biomedicine. Journal of Physics D: Applied Physics 2003, 36 (13), R167-R181.