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  • Affinity Isolation and Purification Matricies

    Protein ExpressionReturn to Magnetic Matrices & Racks

    Separation Racks

    NEB Magnetic Separation Racks are designed to be used for small-scale separations using magnetic particles. Strong neodymium rare earth magnets permit rapid separation of magnetic particles from suspension. Multiple separations can be done using standard sized microcentrifuge tubes with the 6- and 12-Tube Magnetic Separation Racks, and 50 ml tubes with the 50 ml Magnetic Separation Rack.

    Magnetic Protein Purification

    Chitin Magnetic Beads (NEB #E8036): An affinity matrix for small-scale isolation of target proteins fused to a chitin binding domain (CBD) from cell culture supernatants (1). Chitin beads have been prepared having a magnetite core and can be regenerated without loss of binding capacity. Immobilized fusion proteins can be used in subsequent experiments to capture target proteins from crude cell lysates that interact with the immobilized fusion protein.

    Amylose Magnetic Beads (NEB #E8035): An affinity matrix for small-scale isolation and purification of maltose binding protein (MBP) fusion proteins from cell culture supernatants. Amylose is covalently coupled to a paramagnetic particle through a linkage that is stable and leak resistant over a wide pH range. Immobilized fusion proteins can be used in subsequent experiments to capture target proteins from crude cell lysates that may interact with immobilized fusion protein.

    Anti-MBP Magnetic Beads (NEB #E8037): An affinity matrix for small-scale isolation and purification of maltose binding protein (MBP) fusion proteins from cell culture supernatants. Monoclonal anti-MBP is covalently coupled to a nonporous paramagnetic particle through a linkage that is stable and leak resistant over a wide pH range.

    Immunomagnetic Isolation

    Protein A (NEB #S1425) and Protein G (NEB #S1430) Magnetic Beads are affinity matrices for the small-scale isolation and purification of immunoglobulins from many mammalian species including human, rabbit and mouse (1). Truncated recombinant forms of Protein A and Protein G are covalently coupled to a nonporous paramagnetic particle. These truncated proteins exhibit higher unit binding for the IgG Fc region and lower non-specific binding. The affinity of these proteins for IgG varies with species and subclasses of IgG within a species.

    The proteins are coupled through a linkage that is stable and leak resistant over a wide pH range. This permits the immunomagnetic purification of IgG from ascites, serum or cell culture supernatants; after which, the matrix can be re-generated without loss of binding capacity. Protein A and Protein G magnetic beads can also be used to immunoprecipitate target proteins from crude cell lysates using selected primary antibodies. Specific antibodies can be chemically cross-linked to the Protein A coated surface to create a reusable immunprecipitation bead, avoiding co-elution of antibody with target antigen (2,3).

    Immobilized Secondary Antibodies

    Goat Anti-Rabbit IgG Magnetic Beads (NEB #S1432): An affinity matrix for the small-scale immunomagnetic separation and purification of rabbit IgG's. Goat Anti-Rabbit IgG is covalently coupled to a nonporous paramagnetic particle. This secondary antibody binds the heavy chain of all rabbit IgG subclasses and is suitable for immunoassays that employ a rabbit IgG primary polyclonal antibody. Cell separations and sorting can be accomplished using rabbit IgG antibody to defined cell surface antigens.

    Goat Anti-Mouse IgG Magnetic Beads (NEB #S1431): An affinity matrix for the small-scale immunomagnetic separation and purification of mouse IgG's. Anti-Mouse IgG is covalently coupled to a nonporous paramagnetic particle. This secondary antibody binds the heavy chain of mouse IgG and is suitable for immunoassays that employ a mouse IgG primary monoclonal antibody. Cell separations and sorting can be accomplished using a mouse IgG antibody to defined cell surface antigens.

    Purification of Biotin-labeled Substrates

    Streptavidin magnetic beads (NEB #S1420) are 1 µm superparamagnetic particles covalently coupled to a highly pure form of streptavidin. The beads can be used to capture biotin labeled substrates including antigens, antibodies and nucleic acids (4,5). Captured substrates are useful as ligands in subsequent experiments including mRNA isolation and the capture of primary or secondary antibodies.

    References

    1. Harlow, E. and Lane, D. (1988). Bacterial Cell Wall Proteins that Bind Antibodies. Antibodies: A Laboratory Manual, (pp. 615–619). Cold Spring Harbor: Cold Spring Harbor Laboratory Press.
    2. Schneider, C. et al. (1982) J. Biol. Chem. 257, 10766. PMID: 6955305
    3. Sisson, T.H. Caster, C.W. (1990) Immunol. Methods, 127, 215. PMID: 2313100
    4. He, M. and Taussig, M. (1997) Nucleic Acids Res. 25, 5132–5134. PMID: 9396828
    5. Cuddy, K. et al. (1993) Nucleic Acids Res. 21, 2281. PMID: 7684840