Protein A/G Magnetic Co-IP/IP Kit

Immunoprecipitation (IP) or co-immunoprecipitation (Co-IP) is a common technique to study proteins or protein-protein interactions (PPIs), through the use of specific antibodies and antibody-binding media (such as Protein A/G magnetic beads, etc.), Alternatively, a medium conjugated to a specific antibody (e.g., agarose gel or magnetic beads) can be used directly to separate the protein of interest from complex samples and can then be used for SDS-PAGE or mass spectrometry analysis.

This product is a classic magnetic bead-based IP/Co-IP kit, which contains high-quality Protein A/G magnetic beads and the necessary reagents for immunoprecipitation that have been optimized and validated, making IP or Co-IP experiments simpler, more convenient, and more efficient. Protein A/G can be specifically bound to the Fc terminal of the user's own specific antibody, and after a certain period of incubation, the Protein A/G magnetic bead-antibody mixture (beads-Ab complex) is formed, and then the sample is added, and the sample can be specifically recognized by the Fab end of the antibody to form the Protein A/G magnetic bead-antibody-antigen immune complex (Beads-Ab-Ag complex). The immune complexes are washed to remove unbound proteins, and then the bound immune complexes are eluted from the beads for subsequent detection using methods such as acidic eluent or SDS-PAGE loading buffer.

Protein A is a cell wall surface protein found in Staphylococcus aureus with a molecular weight of 42 kDa, which specifically binds to the Fc region of mammalian immunoglobulin (Ig) and also to the Fab region of the human VH3 family. Protein G is an immunoglobulin-binding protein expressed by Streptococcal bacteria, which specifically binds to the Fc region of mammalian immunoglobulin (Ig). This product is a modified recombinant Protein A (25 kDa) and Protein G (25 kDa), covalently coupled to nanoscale amino magnetic beads, and only the amino acid sequences related to the Fc-terminal binding of IgG are retained, and the sequences that may cause non-specific binding except the binding site are removed, so as to effectively reduce non-specific binding.

Protein A beads can specifically bind to the corresponding antibodies, such as human IgG1, IgG2, IgG4, mouse IgG2a, IgG2b and rabbit IgG, and each Protein A molecule can bind to 5 IgG molecules, while Protein G beads can bind antibodies to human IgG1, IgG2, IgG3, IgG4, mouse IgG1, IgG2a, IgG2b , IgG3, rat IgG1, IgG2a, IgG2b, IgG2c, rabbit, goat polyclonal antibodies, etc., and each Protein G molecule can bind 3 IgG molecules. Protein A/G beads are mainly used for immunoprecipitation (IP), co-immunoprecipitation (Co-IP) or chromatin immunoprecipitation (Ch-IP), as well as for the purification of antibodies in samples such as serum, cell culture supernatant or ascites fluid.

Protein A/G beads are a 1:1 ratio of Protein A beads to Protein G beads, which has a number of significant advantages. First, a high content and binding specificity of the conjugated antibody can be achieved. Compared with traditional Protein A/G agarose gels, this product has a smaller pore size, which is less prone to non-specific adsorption and has a high binding capacity. 1 mL of magnetic bead suspension contains approximately 10 mg of magnetic beads and no less than 0.6 mg of recombinant Protein A/G, which can bind no less than 0.7 mg of Human IgG, and the specific maximum binding amount depends on the type of antibody and the target protein. For experiments, 10–20 μL of Protein A/G beads suspension is typically used for 500 μL samples for efficient immunoprecipitation. Second, ultra-rapid binding of antibodies or antibody complexes can be achieved. Protein A/G beads (~200 nm) facilitate fast and efficient binding of magnetic beads to antibodies or antibody complexes due to their large specific surface area. The adsorption process of the antibody or its complexes can be completed within 10 minutes, and the immunoprecipitation of the target protein can be completed within 30 minutes. Shortening the operation time can effectively avoid the degradation or denaturation of the target protein during long-term operation, and fully ensure the activity of the target protein. Due to the magnetic separation, IP and Co-IP can be performed with up to 40% less time per session compared to agarose gels. Finally, a variety of methods can be used for elution. Depending on the structure of the protein of interest, its biological function, and the design requirements of subsequent applications, it can be eluted using a variety of eluents such as acidic solutions, SDS-PAGE loading buffers, or competitive peptides.