Protein Protocols Protein Protocols

Protein A or Protein G 993
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Purification of IgG Using Protein A or Protein G
Mark Page and Robin Thorpe
1. Introduction
Some strains of Staphylococcus aureus synthesize protein A, a group-specific ligand
that binds to the Fc region of IgG from many species (1,2). Protein A does not bind all
subclasses of IgG, e.g., human IgG 3, mouse IgG 3, sheep IgG 1, and some subclasses
bind only weakly, e.g., mouse IgG 1. For some species, IgG does not bind to protein A
at all, e.g., rat, chicken, goat, and some MAbs show abnormal affinity for the protein.
These properties make the use of protein A for IgG purification limited in certain
cases, although it can be used to an advantage in separating IgG subclasses from mouse
serum (3). Protein G (derived from groups C and G Streptococci) also binds to IgG Fc
with some differences in species specificity from protein A. Protein G binds to IgG of
most species, including rat and goat, and recognizes most subclasses (including human
IgG 3 and mouse IgG 1), but has a lower binding capacity. Protein G also has a high affinity
for albumin, although recombinant DNA forms now exist in which the albumin-binding
site has been spliced out, and are therefore very useful for affinity chromatography. Other
streptococcal immunoglobulin binding proteins are protein H (binds IgG Fc), protein B,
which binds IgA and protein Arp, which binds IgG & IgA. These are not generally
available for immunochemical use.
Another bacterial IgG-binding protein (protein L) has been identified (4). Derived
from Peptostreptococcus magnus, it binds to some κ (but not λ) chains. Furthermore,
protein L binds to only some light-chain subtypes, although immunoglobulins from
many species are recognized (5,6).
Finally, hybrid molecules produced by recombinant DNA procedures, comprising
the appropriate regions of IgG-binding proteins (e.g., protein L/G, protein L/A) also
have considerable scope in immunochemical techniques. These proteins are therefore
very useful in the purification of IgG by affinity chromatography. Columns are commercially
available (MabTrap G II, Pharmacia, Uppsala, Sweden) or can be prepared
in the laboratory. The product of this method is of high purity and is useful for most
immunochemical procedures including affinity chromatography and conjugation with
radioisotopes, enzymes, biotin, and so forth.
From: The Protein Protocols Handbook, 2nd Edition
Edited by: J. M. Walker © Humana Press Inc., Totowa, NJ
993