Appearance and Formation Analysis of Multimers in High Concentration Antibodies

Aggregation of antibodies, especially multimers, is a problem that puzzles scientific and industrial fields, and it is more serious in high concentration antibodies. We found that the high concentration antibody solution with rich multimers has a deeper color, which was confirmed by SEC–HPLC, DLS, and SEC–DLS methods. After that, the reasons for formation of multimers were analyzed. LC–MS was used for complete molecular weight analysis, which showed that the multimer sample could not be ionized, indicating a complex structure of multimers. ICP–MS was carried out to analyze the elements in the multimers, and revealed an important message that iron ion in the multimers was significantly higher. On further analysis of the interaction between iron ions and antibody molecules, the results showed that iron ions (Fe(III)) and antibody monomers irreversibly formed multimers under acidic conditions. This study provided useful insights for optimizing the development of antibody drug substances.