Use of modified chitosan macrospheres in the selective removal of immunoglobulins

A solution of chitosan in acetic acid was atomized into sodium hydroxide solution to yield chitosan beads that were 400-600 μm in diameter and with a solids content of 3.5%. Chitosan beads were modified to include a spacer arm and end-capped with a carboxyethyl-group containing anionic ligand to generate a support for use in bioseparations. The ligand modified chitosan beads will be further referred to as LMCB. The influence of pH, salt concentration, and chaotropic salts on the binding of immunoglobulins (Igs) to LMCB has been studied. LMCB supports allowed the separation of monoclonal antibodies (Mab) from cell culture supernatants. In addition, different Mab sub-species (IgG ₁, IgG_2a, IgG_2b, and IgG₃) were also retained from cell culture supernatants rich in the same and further eluted to yield purified Mab. Elution was possible under mild conditions with a step salt gradient. Overall protein recoveries in the range of 90-95% were obtained with cell culture supernatant. Purity of products obtained form a single chromatographic step was estimated to be greater than 98%.