Defined ingredients for defined biology
Cell culture media attempts to recreate the physiologic milieu of cells in their native environment. First attempts were made using blood products, salts and essential nutrients. Over time, more specific mixtures were developed, yet a reliance on blood products often remained, resulting in variability and the possibility of transmitting pathogens. To address this issue and completely eliminate animal ingredients, researchers have developed media that is constituted exclusively from defined ingredients, including recombinant proteins, to create defined media. This is of particular importance as cells are used to produce therapeutic proteins, such as hormones or antibodies, or the cells themselves become the therapy, such as in CAR-T. Here, regulatory oversight and manufacturing quality requires all ingredients to be known provenance.
A particularly sensitive and challenging branch of cell biology is the domain of stem cell culture. Here, undifferentiated cells must be maintained in a toti- or pluri-potent state for many passages to remain both mitotic and undifferentiated and yet responsive to later directed differentiation cues. Cell culture media must be precisely formulated to maintain the cells in this undifferentiated state. In contrast, the differentiation of stem cells into specific cell types, such as cardiomyocytes or neural lineages, requires specific media and growth factors that recapitulate as faithfully as possible the specific genetic programs that occur in the embryonic animal to create the cells that comprise the heart and nervous system, respectively.
HiDef-B8: animal-free, serum-free, highly optimized pluripotency medium
Robust and well-characterized protocols are of utmost importance when culturing hiPSCs for downstream applications. High pluripotency, genetic stability, large-scale production and maintained function all must be kept in mind for hiPSCs. Therefore, much focus has been placed on optimizing cell culture media, growth factors, small molecules and other additives to maintain hiPSCs for large volumes and time scales with high efficiency, low-cost and robustness. This has led to the development of numerous culture media recipes since the early 2000s for hESCs and onwards for hiPSCs. This has included the use of KSR and FGF2 for feeder-free media and Activin A, xeno-free media development, and the addition of Rho-associated, coiled-coil containing protein kinase (ROCK) inhibitors and vitronectin, among others. These advances led to many variants of hiPSC media over the years, including DC-HAIF, hESF9, TeSR, CDM-BSA, E8, FTDA, and iDEAL.
While much has been learned along the way, many of these media fail to satisfy the full needs of hiPSC research, including robustness over multiple passages and experimental needs, low cost, ease of production, weekend-free media changes and fully defined components. Such traits are critical for robust, consistent and affordable research in the hiPSC space. To meet these demands in a scalable manner, Defined Bioscience introduces HiDef-B8, a robust, weekend-free, highly validated iPSC medium originally developed by the Paul Burridge lab at Northwestern University. This formulation from Defined Bioscience requires the addition of only a single vial of contents to a bottle of standard DMEM/F12 medium, and maintains iPSCs in a pluripotent state past 100 passages, as confirmed by flow cytometry using standard pluripotency markers.