# An Edible, Decellularized Plant Derived Cell Carrier for Lab Grown Meat
## 4 Discussion
Media was changed frequently; however, at no point were the scaffolds and cells agitated or suspended in the culture media, as they would be in a large suspension bioreactor. In this study, cells were inoculated directly into an environment already containing culture medium and unsupported scaffolds, and the environment was intermittently agitated to generate random flow fields that suspended the reactor contents. In this dynamic environment cells adhered to and remained viable on the decellularized plant-based scaffolds, and ultimately suggested that traditional cell inoculation strategies associated with microcarrier culture applied to primary bovine satellite cells and decellularized plant-based carriers. This method removed the time and user intensive process of cell seeding, which is important in reducing costs in the cellular agriculture industry. Suspension style reactors generate fluid flow fields that suspend cell-populated microarrays in the interest of maximizing the number of cells cultured per unit volume. It is important to minimize the fluidic stresses imparted upon cells by such flow fields, typically generated via stirring impeller, or rocking platform, as such stresses have been showed to impact mammalian cell expansion [27]. Computational fluid dynamics was used to calculate the patterns of fluid flow within such reactors so that cell engineers could predict the stresses that their cell carriers might encounter during expansion [28]. Data pertaining to the size, shape, and density of the microcarriers was paramount for predicting the minimum carrier suspension criteria, as well as the mechanics by which the carrier passed through the media during suspension [29]. Microcarriers are currently fabricated with a density slightly greater than that of the media within which they are suspended. This ensures that microcarriers in a static environment will settle, allowing for initial cell attachment during inoculation, and can then be suspended with minimal energy input. The decellularized floret scaffolds were found to have a density of approximately 1.01 g/cm\({}^{3}\). This density measurement was comparable to microcarriers previously suggested for lab grown meat, and those historically used for suspension culture of anchorage-dependent cells for biomedical applications [9]. These materials, comprised of edible polysaccharides and polypeptides varied in density between 1.03 g/cm\({}^{2}\) and 1.04 g/cm\({}^{2}\)[30; 31; 32; 33] (Figure 7). Cell culture media containing 10% FBS has been reported at 1.007 g/m\({}^{3}\) and decreased with lesser concentrations of FBS [34]. Such similar density values implied that the suspension criteria for cells cultured on decellularized florets would be minimal when suspending the cell carriers. These suspension criteria served as the conditional basis for computational fluid dynamics and generated the bioreactor's velocity gradient, which would ultimately be used to identify the fluid flow profile of the bioreactor. The size and shape of the carrier were key variables in predicting the stresses cells were expected to encounter [29]. A larger carrier, must displace greater fluid volumes as they pass around it, and thus will encounter greater stress magnitudes. The ability to sort our decellularized floret carriers by size, as demonstrated, might be a means of minimizing stress-induced cell damage. It was interesting to note that the cross-sectional images generated via histological staining revealed relatively large spaces within the bull of the floret scaffold. Porous microcarriers have been developed in the past as a means of protecting cells that have infiltrated the bead from shear stresses that they might encounter [10]. It is recommended that the porosity of decellularized floret microcarriers, and of any future developed decellularize plant-derived microcarriers, is examined. Additionally, the surface of a more eccentric particle is expected to encounter greater shear stresses than normal stresses as fluid volumes pass around it.