Pitfalls in Early Bioprocess Development Using Shake Flask Cultivations

For about 100 years, the shake flask has been established for biotechnological cultivations as one of the most important cultivation systems in early process development. Its appeal lies in its simple handling and highly versatile application for a wide range of cell types-from bacteria to mammalian cells. In recent decades, extensive research has been conducted on the shake flask, to not perform processes blindly but to gain a deeper understanding of the various process parameters, phenomena, and their impact on the process. Although the characterization of the shake flask is now well-established in literature, many publications show that this knowledge is often inadequately applied. Therefore, this review provides an overview of the current state of knowledge on various topics related to the shake flask. We first present the key process parameters and their influence on different physical phenomena, such as power input, the largely unknown in-phase/out-of-phase phenomenon, as well as temperature and mass transfer. Then, the most common online monitoring systems that have been established for shake flasks are discussed. Finally, various pitfalls that often arise from inadequate knowledge of handling shake flask cultivations are discussed and guidance on how to avoid them is provided.