Magnetic-activated cell sorting
Encyclopedia
Magnetic-activated cell sorting (MACS) is a trademark name for a method for separation of various cell
populations depending on their surface antigens (CD molecules
). The term MACS is a registered trademark of Miltenyi Biotec.
Magnetic cell separation using antibodies is also possible using monosized magnetic beads (Dynabeads
) from Invitrogen (now part of Life Technologies). The Dynabeads technology is not column based - instead these magnetic beads with attached cells enjoy liquid phase kinetics in a sample tube, and the cells are isolated by placing the tube on a magnetic rack.
coated with antibodies
against a particular surface antigen. This causes the cells expressing this antigen to attach to the magnetic nanoparticles. Afterwards the cell solution is transferred on a column placed in a strong magnetic field. In this step, the cells attached to the nanoparticles(expressing the antigen) stay on the column, while other cells (not expressing the antigen) flow through. With this method, the cells can be separated positively or negatively with respect to the particular antigen(s).
lymphocyte
s.
-conjugated antibodies are much more prevalent, it is possible to use magnetic nanoparticles coated with anti-fluorochrome antibodies. They are incubated with the fluorescent-labelled antibodies against the antigen of interest and may thus serve for cell separation with respect to the antigen.
Cell (biology)
The cell is the basic structural and functional unit of all known living organisms. It is the smallest unit of life that is classified as a living thing, and is often called the building block of life. The Alberts text discusses how the "cellular building blocks" move to shape developing embryos....
populations depending on their surface antigens (CD molecules
Cluster of differentiation
The cluster of differentiation is a protocol used for the identification and investigation of cell surface molecules present on white blood cells, providing targets for immunophenotyping of cells...
). The term MACS is a registered trademark of Miltenyi Biotec.
Magnetic cell separation using antibodies is also possible using monosized magnetic beads (Dynabeads
Dynabeads
Dynabeads are superparamagnetic, monosized and spherical polymer particles with a consistent, defined surface for the adsorption or coupling of various bioreactive molecules or cells....
) from Invitrogen (now part of Life Technologies). The Dynabeads technology is not column based - instead these magnetic beads with attached cells enjoy liquid phase kinetics in a sample tube, and the cells are isolated by placing the tube on a magnetic rack.
Procedure
The MACS method allows cells to be separated by incubating with magnetic nanoparticlesNanoparticle
In nanotechnology, a particle is defined as a small object that behaves as a whole unit in terms of its transport and properties. Particles are further classified according to size : in terms of diameter, coarse particles cover a range between 10,000 and 2,500 nanometers. Fine particles are sized...
coated with antibodies
Antibody
An antibody, also known as an immunoglobulin, is a large Y-shaped protein used by the immune system to identify and neutralize foreign objects such as bacteria and viruses. The antibody recognizes a unique part of the foreign target, termed an antigen...
against a particular surface antigen. This causes the cells expressing this antigen to attach to the magnetic nanoparticles. Afterwards the cell solution is transferred on a column placed in a strong magnetic field. In this step, the cells attached to the nanoparticles(expressing the antigen) stay on the column, while other cells (not expressing the antigen) flow through. With this method, the cells can be separated positively or negatively with respect to the particular antigen(s).
Positive selection
In positive selection the cells expressing the antigen(s) of interest, which attached to the magnetic column, are washed out to a separate vessel, after removing the column from the magnetic field. This method is useful for isolation of a particular cell type, for instance CD4CD4
CD4 is a glycoprotein expressed on the surface of T helper cells, monocytes, macrophages, and dendritic cells. It was discovered in the late 1970s and was originally known as leu-3 and T4 before being named CD4 in 1984...
lymphocyte
Lymphocyte
A lymphocyte is a type of white blood cell in the vertebrate immune system.Under the microscope, lymphocytes can be divided into large lymphocytes and small lymphocytes. Large granular lymphocytes include natural killer cells...
s.
Negative selection
In negative selection the antibody used is against surface antigen(s) which are known to be present on cells that are not of interest. After administration of the cells/magnetic nanoparticles solution onto the column the cells expressing these antigens bind to the column and fraction that goes through is collected, as it contains almost no cells with undesired antigens.Modifications
Magnetic nanoparticles conjugated to an antibody against an antigen of interest are not always available, but there is a way to circumvent it. Since fluorophoreFluorophore
A fluorophore, in analogy to a chromophore, is a component of a molecule which causes a molecule to be fluorescent. It is a functional group in a molecule which will absorb energy of a specific wavelength and re-emit energy at a different wavelength...
-conjugated antibodies are much more prevalent, it is possible to use magnetic nanoparticles coated with anti-fluorochrome antibodies. They are incubated with the fluorescent-labelled antibodies against the antigen of interest and may thus serve for cell separation with respect to the antigen.