Challenge
One of our sponsors requested immunoassay development and validation to measure a humanized therapeutic monoclonal antibody (mAb) fragment (Fab) that inhibits a human growth factor involved in the pathogenesis of a common degenerative disease of aging.
Antibody fragments offer numerous advantages over full-length antibodies in terms of production cost, increased penetration into the target tissue, and reduced potential for inappropriate activation of immune effector mechanisms. However, because antibody fragments lack many common domains typically targeted (e.g., Fc regions) by immunoassay reagents for binding and detection, they may also present unique challenges for bioassay design, potentially impacting sensitivity since fewer antigens/antibodies can bind to each fragment.
We could try to optimize the concentration of coating and detection reagents to improve sensitivity, but this strategy carries the risk of increasing background noise due to specific and nonspecific interactions within the various assay components (e.g., the therapeutic molecule, the blocking agents, and/or the coating antigen). The potential for off-target specific interactions is especially concerning when the matrix belongs to a closely related species, in this case a non-human primate.