title: Purification of monoclonal antibodies by hydrophobic interaction chromatography under no-salt conditions.
abstract: Hydrophobic interaction chromatography (HIC) is commonly used as a polishing step in monoclonal antibody purification processes. HIC offers an orthogonal selectivity to ion exchange chromatography and can be an effective step for aggregate clearance and host cell protein reduction. HIC, however, suffers from the limitation of use of high concentrations of kosmotropic salts to achieve the desired separation. These salts often pose a disposal concern in manufacturing facilities and at times can cause precipitation of the product. Here, we report an unconventional way of operating HIC in the flowthrough (FT) mode with no kosmotropic salt in the mobile phase. A very hydrophobic resin is selected as the stationary phase and the pH of the mobile phase is modulated to achieve the required selectivity. Under the pH conditions tested (pH 6.0 and below), antibodies typically become positively charged, which has an effect on its polarity and overall surface hydrophobicity. Optimum pH conditions were chosen under which the antibody product of interest flowed through while impurities such as aggregates and host cell proteins bound to the column. This strategy was tested with a panel of antibodies with varying pI and surface hydrophobicity. Performance was comparable to that observed using conventional HIC conditions with high salt.
authors.affiliation: Department of Process Biochemistry, Biogen Idec; Research Triangle Park, NC USA.
authors.forename: Sanchayita
authors.initials: S
authors.lastname: Ghose
authors.forename: Yinying
authors.initials: Y
authors.lastname: Tao
authors.forename: Lynn
authors.initials: L
authors.lastname: Conley
authors.forename: Douglas
authors.initials: D
authors.lastname: Cecchini
mesh_terms: D000645:Ammonium Sulfate / Q000737:chemistry; D000911:Antibodies, Monoclonal / Q000737:chemistry* / Q000302:isolation & purification*; D002845:Chromatography / Q000379:methods*; D002951:Citrates / Q000737:chemistry; D006863:Hydrogen-Ion Concentration; D057927:Hydrophobic and Hydrophilic Interactions; D008970:Molecular Weight; D010710:Phosphates / Q000737:chemistry; D017680:Potassium Compounds / Q000737:chemistry; D015203:Reproducibility of Results; D012492:Salts / Q000737:chemistry*; D000077559:Sodium Citrate
chemical_list: D000911:Antibodies, Monoclonal; D002951:Citrates; D010710:Phosphates; D017680:Potassium Compounds; D012492:Salts; D000077559:Sodium Citrate; C013216:potassium phosphate; D000645:Ammonium Sulfate
keywords: HIC; aggregates; flowthrough; monoclonal antibodies; no salt