Cell culture growth surface

The use of buccal cell cultures for assessing the permeability of the buccal mucosa has attracted recent attention (see Chap. 7 for a more extensive summary). In order to culture buccal epithelial cells, the cells must be harvested from an appropriate source and cultured under specific conditions using an appropriate growth medium, temperature, and humidity [46], Cell cultures have been successfully grown from hamster cheek pouch. These cultured cells, however, did not differentiate to form a complete keratinized surface as seen in the normal hamster cheek pouch, and they consequently displayed a greater permeability to compounds when compared with keratinized hamster cheek pouch mucosa [134], Therefore, the cultured hamster cheek cells more closely mimicked the human buccal mucosa due to their lack of keratinization, and so this may be an appropriate model for predicting permeability through the human buccal mucosa. 

Cell culture systems must provide the physiological conditions for cell survival and proliferation. In vitro, animal cell growth is dependent on several factors, such as pH, temperature, osmolality, gas concentration (oxygen and CO2), available surface substrata, and state of the cells at inoculation (Freshney, 2005). Other factors that impact the culture are medium composition, which can differ extensively between cell lines and is discussed in detail in Chapter 5, as well as susceptibility to hydrodynamic stress, as discussed in Chapter 7. 

As mentioned in Chapter 9, since production scale-up is related to the increase of cell culture surface for adherent cells, consideration must be given to the relationship between the surface area available for cell growth and the bioreactor volume (Kent and Mutharasani, 1992). 

Fig. 3.2(a). Vessels in which cells may be cultured. At the top left are three sizes of flask (125, 75 and 25 cm2) and at the bottom left three sizes of dish (9.5 and 3 cm diameter). To the right are shown 6, 12 and 24-well tissue culture trays and a 96-well microtitre plate. At the bottom right is an 8-chamber culture slide, (b) Millicell inserts for 6-well TC trays provide a semi-permeable growth surface. 

The dilute cell suspension may be plated out on special Petri dishes whose growth surface is made of a membrane (Petriperm Appendix 3). This membrane allows ready diffusion of gases in and out of the dish from below and is also readily cut into pieces. In this way pieces containing single colonies may be cut out and transferred to the wells in a tissue culture tray (cf. broken cover-slips). 

Cells are grown either in suspension in a free or immobilized form 102), or by adherence to a solid surface 100). Materials used for promoting surface-dependent cell growth are glasses, metals, plastics, carbohydrate polymers etc. the media used contain substances such as blood plasma, amniotic fluids, tissue extracts, etc.103). Recent developments in animal cell culture are aimed at the improvement of strains and culture techniques, medium optimization, and scale-up. In contrast to plant cell culture, animal cell culture has already found its technical application. Large-scale... 

Other primary cells, like cardiomyocytes or chondrocytes, can rapidly undergo de-differentiation to fibroblast-like cells when plated on a two-dimensional (flat) surface. In this case, three-dimensional matrices can help maintain the original features. BioLevitator by Hamilton Robotics, 3D Insert by 3dbiotek, and 3D cell culture by Invitrogen are only some of the products available to facilitate the correct growth and morphology of particular primary cell types. 

Biopharmaceuticals that are industrially produced by microbial (bacterial or yeast) fermentation include insulin, human growth hormone, hepatitis B surface antigen vaccine, alpha interferon, beta interferon, gamma interferon, granulocyte colony stimulating factor, and interleukin-2. Table 30.8 lists some of the major biopharma products, some of which are produced by mammalian cell culture. 

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