Technical Scale in Biomolecular Crystallization

In recent years, the scalability of biomolecular crystallization processes has garnered significant attention due to its advantages in enhancing product quality and reducing manufacturing costs compared to the traditional purification methods. Unlike the crystallization process of small molecules, biomolecular crystallization lacks a comprehensive theory or guidelines, often relying on modified crystallization theories and empirical approaches. It is challenging to guide protein molecules assembling into ordered crystal lattice due to the huge molecular size and complexity of the structure, and protein crystals are more delicate and fragile than conventional small organic molecular crystals. Furthermore, proteins often exhibit limited stability within specific environmental conditions, such as varying pH and temperature ranges. Consequently, many standard methods employed for small-molecule crystallization are not applicable to biomolecular crystallization. Most of the bimolecular crystallization research are focused on tiny scale, while more research on biomolecular crystallization at various larger technical scales has emerged over time. The scaling up mechenism from the tiny scale to the large scale is critical to widely apply the biomolecular crystallization. This chapter begins by summarizing the different methods used in biomolecular crystallization. Then, biomolecular crystallization is reviewed from nL to μL scale (e.g., hanging drop or sitting drop) to mL scale, such as in small batch or microfluidic crystallizer till hundred mL or L scale, such as standard stirred crystallizer. Finally, it explores different crystallization scales with corresponding process monitoring and analytical methods for the crystallization kinetics.