Cation exchange chromatography is commonly used as a polishing step in the purification of monoclonal antibodies. Cation exchange resins, however, typically have some lot-to-lot variability, which may result in a visually different elution profile, depending on the conditions used in the chromatography step. Product elution volumes also may be significantly different, depending on the product collection criteria. Despite this, there are no significant differences in the clearance of impurities. Differences in product volume, however, could lead to differences in the pH and conductivity of the product that could have an impact on the subsequent unit operation. This paper will present a case study that shows how the lot-to-lot variability of a cation exchange resin impacts its process performance and the performance of the subsequent process step. The study found that differences in product volume and elution profile were more prominent when resin lots containing larger percentages of smaller particles were used.
In addition to large elution volumes, significant visual differences in shape of elution profile were observed. These differences included a flat portion followed by a trailing portion on the descending part of the peak, instead of a smooth and relatively rapid fall.
To determine the root cause of the unusually wide elution peaks, bench-scale cation exchange experiments were performed on four columns with unused resins. The first column was packed with lot A, the lot of resin that was used for all the initial cation exchange development work. The remaining three columns were packed with lots B, C, and D, respectively.
MATERIALS AND METHODS
Some details relating to materials and methods could not be included because of confidentiality concerns.
Column Packing and Evaluation
Four 1.1-cm diameter Millipore Vanguard columns were each packed with a different lot of cation exchange resin to a height of 20±2 cm. Equilibration buffer was applied to each column. After the columns were equilibrated, approximately 0.25 mL (1.3% CV) M NaCl was injected. The conductivity of the column effluent was monitored. Column performance was evaluated by calculating the asymmetry and the height of a theoretical plate (HETP) from the NaCl chromatogram. Following this, the columns containing lots B, C, and D were also evaluated with a pulse acetone injection. The resulting signal was measured with UV at 280 nm.