Cell propagation Suspension cells

Figures 9.2 and 9.3 show schemes that illustrate inoculum development from the cryotubes to production scale for suspension and adherent cells, respectively. In these hypothetical process schemes, the expression production bioreactor is used arbitrarily for any of the types of bioreactor presented in the next section of this chapter. In general, different flasks and several intermediate bioreactors are used for cell propagation to reach the quantity of cells necessary to inoculate the production bioreactor. The number of propagation steps is a function of the final scale of the production bioreactor.

Hypothetical example of the propagation of suspension cells, using stirred-tank bioreactors in the intermediate and final stages. The amount of cells usually obtained at the end of each step is also indicated. 

Himmelfarb P, Thayer PS, Martin HE (1969), Spin filter culture the propagation of mammalian cells in suspension, Science 164 555-557. 

Katinger HWD, Scheirer W Krdmer E (1979) Bubble column reactor for mass propagation of animal cells in suspension culture. German Chemical Engineering 2 31. 

This procedure describes the preparation of nuclear and cytoplasmic extracts from cells grown in suspension (2-201) and is essentially as described by Dignam et al.1 and modified by Jamison and Garcia-Bianco.2 A procedure for small-scale preparation of extracts from HeLa cells grown as monolayer has been described by Lee and Green.3 The latter procedure is recommended when the cell material is scarce, if radioactively labelled extracts are made, if several extracts from parallel cultures, or if expensive growth conditions are necessary for cell propagation. 

Most El-deficient recombinant adenoviruses are propagated in the HEK-293 and PER.C6 cell lines (and derivatives thereof) as discussed in Section 10.1.2.3. The generation of RCAs is minimal in PER.C6 but remains a concern for HEK-293. Both cell lines have been documented for GMP manufacturing [110]. The production of these cells can be accomplished by a variety of methods, which depend on whether adherent or suspension cell lines are being used. 

Steiner D, Khaner H, Cohen M, Even-Ram S, GU Y, Itsykson P, Turetsky T, Idelson M, Aizenman E, Ram R, Berman-Zaken Y, Reubinoff B. (2010) Derivation, propagation and controlled differentiation of human embryonic stem cells in suspension. Nat. BiotechnoL, 28(4) 361-364. 

Several attenuated strains have been developed for use in vaccine preparations. The most commonly used is the Jeryl Linn strain, which is propagated in chick embryo cell culture. This vaccine has been administered to well over 50 million people worldwide and, typically, results in seroconversion rates of over 97 per cent. The Sabin (oral poliomyelitis) vaccine consists of an aqueous suspension of poliomyelitis virus, usually grown in cultures of monkey kidney tissue. It contains approximately 1 million particles of poliomyelitis strains 1,2 or 3 or a combination of all three strains. 

The BHK-21 (baby hamster kidney) cell line consists of adherent fibroblasts, that can also be adapted to suspension culture, and was isolated from five 1-day-old hamsters (McPherson and Stoker, 1962). These cells are commonly used for virus propagation (polio, rabies, and foot-and-mouth disease) for production of veterinary vaccines. 

In the meantime, the formation of the main alkaloids in C. ipecacuanha under a variety of conditions has been extensively investigated emetine (1) in callus cultures (49) and under the effects of L-tyrosine supplementation (5t)) emetine (1) and cephaeline (2) in Panamanian ipecac (57), in Nicaraguan ipecac (52), in regenerates obtained by clonal propagation (53,54), in tissue cultures (55) and under the effects of exogenous feeding of shikimic acid and L-phenylalanine (55), in cell suspension and excised root cultures (57), in adventitious root cultures (58), and in callus cultures (56,59) and the effects of age and electrokinetic potential (60) ipecoside (7) in the roots (61) and the effect oi Azotobacter, leaf mold, and farmyard manure on alkaloid content (62). In addition, micropropagation systems for C. ipecacuanha have been developed (63-65). 

BY-2 tobacco culture was propagated as previously described. Cells were harvested 1-12 d after sub-culturing, and used for experiments after addition of 10 /tM CLA. LaCla solution (various concentration) was added to cell suspension in glass tubes placed in a CHEM-GLOW Photometer (AMINCO., Silver Spring, MD, USA), and OXB was monitored by CLA-CL, and expressed as relative CL units (rcu) as previously described. Cells were treated with ZnS04 (0.3, 1, 3 mM), 2 min... 

Plant cell and tissue culture methods can be used for the mass propagation of plants. Cells (often from the growing tip) can be grown in a medium containing one hormone to form a callus or a suspension. 

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