Roller flask culture

For roller flask cultures of normal and virus-transformed cells ... 

H. Roller-Flask Cultures of Diploid Cell Lines... 

When employing roller-flask cultures to harvest colchicine-arrested metaphases, calcium is omitted from the medium only during the last day. Colchicine (2.5 X 10 m stock) is added at this time in the preparation of 0.3 ml colchicine per 100 ml medium. After 18-24 hr of treatment, followed by several gentle swirls of the medium, up to 50-55 percent of the cells will detach as metaphases. This amounts to approximately 20 x 10 metaphases per flask per 24 hr. Similarly, calcium must be omitted from the medium when harvesting metaphases for synchrony studies without the aid of colchicine. 

CHO cells should be grown in medium consisting of a-MEM containing 10% fetal calf serum, 100 U/ml penicillin, and 100 /ig/ml streptomycin (complete medium). To provide sufficient cells for each experiment it is most convenient to maintain them in 150-cm tissue culture flasks. However, subsequent preparation of mitotic cells is most easily accomplished using roller bottle cultures. As a rough guide two 950-cm roller bottles should provide a total of about 300-500 p of packed mitotic cells. 

Treatment is normally carried out in 50 ml centrifuge tubes on a roller machine. During the expression time the cells are grown in T75 plastic tissue culture flasks. 

The range of culture flasks and reactor types employed is quite wide, both for suspension and adherent cultures, from small Carrel s or Roux s flasks to roller bottles. Fixed- and fluidized-bed bioreactors, air-lift reactors and even stirred and aerated tanks with capacities up to 15 m3 are common in large plants producing monoclonal antibodies (mAbs) for anticancer therapies (Adams and Weiner, 2005 Griffiths, 1988). 

The basic need for a solid support guides all production choices involving industrial processes for adherent cells. A large variety of vessels has been developed for adherent cell cultures. Petri dishes, Roux bottles, T-flasks, and roller bottles are examples of cell culture vessels with a glass or polystyrene surface. The system of choice is dependent on the seal-ability of multiple steps, as well as the cost of equipment and qualified operators. 

Alternatively, some subunit viral vaccines can be generated by rDNA techniques and expressed in a continuous cell line or insect cells. Recent advances in bio reactor design and operation have improved the successful production of IPV in large-scale bioreactors. However, roller bottles or flasks are still used for most current vaccine production. Development of insect cell culture will allow for very large-scale liquid suspension culture (143). Several vaccine candidates such as gpl60 for HIV and gD protein for herpes have been demonstrated in the insect cell culture system. However, no vaccine has... 

Cell lines L929, AF1-19T. Tissue culture flasks and Roller bottles. Medium Dulbecco s MEM/10% FCS (D/10). 

Because most routine cell types were originally grown on glass, the first commercially available tissue culture surface was modelled after glass chemistry. Conventional tissue culture surfaces therefore are hydrophilic and have an oxygenated chemistry and a net-negative surface charge. This chemistry is basically the same whether the treatment process is produced by corona or plasma. This is the routine surface that is commercially available from a number of different suppliers on plastic dishes, flasks, plates and roller bottles. 

Adherent cells can be grown in stationary or microcarrier cultures [116]. Stationary cultures use devices such as flasks, roller bottles, and cell factories that provide wall surface area for cell attachment [117]. Microcarrier cultures, on the other hand, can be implemented in agitated bioreactors [114,116]. These microcarriers are small porous particles (0.2mm) that provide large surface areas for cell attachment and can be suspended in the culture medium by gentle agitation. The culture environment is thus easily controlled and the scale-up is done by increasing the... 

Tissue culture flasks, roller bottles, and filterwares... 

Since the 1960s, surface modification has been used to enable cells to be cultured on normally inhospitable plastic. The marketing of plasma-treated polystyrene microplates, flasks, roller bottles and dishes has allowed cell culture to be carried out quickly and cheaply on disposable base materials, avoiding the need to culture in glass (in vitro) and reducing the possibility of cross-contamination. More recently, the challenges associated with the culture of primary and pluripotent cells and a focus on the reduction or elimination of animal-derived products in cell culture have stimulated the development of novel surface-coated cell culture products. 

Cell seed expansion A series of culture steps is needed to expand the cell seed ampoule (e.g., 5 million cells) to production size (range 10 to 10 cells). For HDC, this is accomplished in steps of a split level of 1 2 or more usually 1 4 through a series of flasks, roller bottles, and possibly cell factories (A/S Nunc). Other cell types are split at a 1 5 to 1 20 ratio. A similar build-up is needed for the virus seed. 

ADC - Flasks, Roller Bottles, Cell Factories or or Microcarrier Culture SUSP -spinners,fermenters, immobilised reactors) ... 

The steps between the cell bank and the production fermentation serve the purpose of expanding the seed volume (biomass) to finally provide sufficient cells to inoculate the main fermenter. Early seed stages, also referred to as precultures, are often run in shake flasks or in T-flasks and roller bottles in the case of cell culture, whereas subsequent stages are performed in stainless steel or single-use bioreactors. The dimensions of the seed bioreactors are adjusted to the biological system, as shown in Table 1.5. 

---