Protein A, Protein G, Protein A/G and Protein L are native and recombinant proteins of microbial origin that bind to mammalian immunoglobulin molecules. These proteins are available in purified, salt-free, lyophilized form, as well as coated in microplates and covalently immobilized to various solid supports. The most popular support for column affinity purification of antibodies is crosslinked beaded agarose, although Thermo Scientific™ UltraLink™ Biosupport (polyacrylamide) and other porous media are also available. Magnetic beads are the most popular support material for immunoprecipitation applications.

General characteristics of Ig binding proteins

  Native Protein A Recombinant Protein A Recombinant Protein G Recombinant Protein A/G Recombinant Protein L
Native source Staphylococcusaureus Staphylococcusaureus Streptococcus N/A Peptostreptococcus magnus
Production source S. aureus E. coli E. coli E. coli E. coli
Molecular weight 46,700 44,600 21,600 50,460 35,800
Apparent mass by SDS-PAGE 42kDa 45kDa 32kDa 40 to 45kDa 36kDa
# binding sites for Ig 5 5 2 4+2 4
Albumin binding site No No No No No
Optimal binding pH 8.2 8.2 5 5 to 8.2 7.5
Ig binding target Fc Fc Fc Fc VL-kappa

The four proteins bind almost exclusively with the IgG class of antibodies, but their binding properties differ among species and subclasses of IgG. Protein A is generally preferred for rabbit, pig, dog and cat IgG. Protein G has better binding capacity for a broader range of mouse and human IgG subclasses (IgG1, IgG2, etc.).

Protein A/G is a recombinant fusion protein that includes the IgG-binding domains of both Protein A and Protein G. Therefore, Protein A/G is ideal for binding the broadest range of IgG subclasses from rabbit, mouse, human and other mammalian samples.

Protein L binds to certain immunoglobulin kappa light chains. Because kappa light chains occur in members of all classes of immunoglobulin (i.e., IgG, IgM, IgA, IgE and IgD), Protein L can purify these different classes of antibody. However, only those antibodies within each class that possess the appropriate kappa light chains will bind. Generally, empirical testing is required to determine if Protein L is effective for purifying a particular antibody.