To investigate structure and function of different monoclonal antibody (MAb) dimers.
MAb dimers were induced by process-related, low pH and UV light stress. Dimers were isolated and purified by chromatography and extensively characterized by biochemical, structural and functional methods.
Highly purified dimer forms were obtained which enabled detailed characterization. Dimers induced by process stress were associated by a single non-covalent interaction site between two Fab domains in a characteristic "bone-like" structure observed in Transmission Electron Microscopy (TEM). These dimers showed reduced potency and antigen binding affinity. Low pH stress generated more stable but also non-covalently associated dimers without chemical alterations in a typical "closed" conformation according to TEM. These dimer species were more compact and more hydrophobic as dimers induced by process stress. They showed bioactivity and antigen binding affinity similar to the native monomer. Light-induced dimers, exhibiting various different conformations, were the most stable dimers with various chemical modifications leading to a broad range in size, charge and hydrophobicity. These dimers fully lost bioactivity and antigen binding affinity.
The use of highly purified MAb dimers and a panel of characterizations methods enabled to obtain a clear picture about molecular architecture and function of dimers.