Purification of anti-glycoconjugate monoclonal antibodies using newly developed porous zirconia particles.

Here, we describe porous zirconia particles (PZPs) optimized for the purification of immunoglobulins. PZPs, with a pore size of approximately 10 nm, were designed to specifically interact with antibodies via surface modification with a phosphate functional group. A simple PZP purification method based on precipitation enabled efficient purification of mouse anti-glycosphingolipid globoside/Gb4Cer monoclonal IgM (κ-light chains) from hybridoma culture supernatants. Over 99% of contaminating proteins were removed by the PZP purification process, and approximately 50% of the IgM was recovered in the purified fraction after eluting the PZP-adsorbed antibodies with 100 mM phosphate buffer. Other IgG3 and IgM monoclonal antibodies that react with Gb4Cer or α2,6-sialyl LacNAc-modified glycoproteins could also be purified using PZPs and elution buffer at concentrations of 100-500 mM. All of the purified antibodies retained their antigen reactivity and specificity, indicating that PZP purification does not affect antibody function. As PZP purification is also suitable for purification of IgM consisting of λ-light chains and IgG derived from other mammalian species, it is expected to be applied to the purification of a variety of antibodies, including anti-glycoconjugate IgMs.