Application of chemically defined medium and high cell-density fermentation to produce domain antibodies (dAbs) in recombinant Escherichia coli

Domain antibodies (dAbs) are the smallest known antigen-binding fragments of antibodies, ranging from 11 -15 kDa, and are the robust variable regions of the heavy and light chains of immunoglobulins.  dAbs are emerging as a relatively new class of medicine. Currently, our dAbs are produced in E. coli using complex media. Product is secreted into the periplasm and released into the culture medium in a spontaneous manner involving cell lysis. The use of complex medium for large (commercial)-scale application is a risk because of its composition complexity. Potential problems include: (1) raw materials lot to lot variability, (2) process performance inconsistency (e.g. timing of spontaneous release), and (3) scale-up challenges. The use of chemically defined medium can overcome and/or reduce those issues. In this presentation, we will discuss the strategies to express dAbs at high concentrations in the E. coli using chemically defined medium and high cell-density fermentation. To ensure the process performance consistency at the large- scale, we developed a new feed and induction strategy to retain the dAbs inside the periplasm, and recover the product through controlled cell lysis using homogenization. Moreover, the highest amount of a soluble dAb product was achieved at 28 g per L of culture which was much higher than that previously seen with complex media using this approach. The approach presented here can be extended to other fermentation processes. It provides a new approach to solve potential pilot and commercial scale issues and to improve process controllability and consistency.