A new purification method for the Fab and F(ab′)2 fragment of 145-2C11, hamster anti-mouse CD3ε antibody

Recombinant protein G has been utilized in the purification of antibodies from various mammalian species based on the interaction of antibodies with protein G. The interaction between immunoglobulin and protein G may not be restricted to the Fc portion of antibodies, as many different F(ab')(2) or Fab fragments can also bind to protein G. I found both Fab and F(ab')(2) of 145-2C11, a hamster anti-mouse CD3 epsilon antibody, bound to the protein G-sepharose, Interestingly, Fab and F(ab')(2) of 145-2C11 did not bind to the protein A-sepharose. The binding of Fab and F(ab')(2) of 145-2C11 to protein G provided a useful method to remove proteases, chopped fragments of the Fc region, and other contaminating proteins. The remaining intact antibody in the protease reaction mixture can be removed by using a protein A-sepharose, because the Fab and F(ab')(2) portions of 145-2C11 did not bind to protein A-sepharose. The specific binding of Fab and F(ab')(2) portions of 145-2C11 to a protein G-sepharose (though not to a protein A-sepharose) and binding of intact 145-2C11 to both protein A- and G-sepharose will be useful in developing an effective purification protocol for Fab and F(ab')(2) portions of 145-2C11.