Increasing the Versatility of Recombinant Antibodies targeting mitotic epitopes
Monoclonal antibodies (mAbs) are indispensable reagents across many basic, translational, and biomedical applications from biomolecule detection to protein purification to medical diagnostics and therapeutics. However, traditional mAbs generated in animals have their drawbacks. Concerns regarding traditionally produced mAbs include reproducibility, incomplete characterization, consistent availability, and cost. In short, good mAbs are hard to find for certain targets and biological processes such as mitosis.
To maintain genomic integrity, proper segregation of chromosomes into the two daughter cells must occur. This process is powered and regulated by the kinetochores, which are structures built at the primary constriction of mitotic chromosomes that attach to spindle microtubules. The primary factor connecting the kinetochores to the microtubules is the kinetochore-associated NDC80 complex.
Advancements in protein sequencing techniques have made it possible to acquire the primary amino acid sequence of mAbs that target the NDC80 complex of the mitotic process. The invariant primary sequences were used for downstream construction and production of sequence-defined recombinant mAbs and antibody fragments, including scFvC, scFv, and Fab, all showing robust binding to the kinetochore target in mitotic cells. Moreover, this methodical approach circumvented the issues listed above, providing a straightforward and accessible strategy for generating low-cost, high-yield preparations of recombinant mAbs and antibody fragments.