Scale Production of Recombinant Proteins

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Lundstrom, K. (2003) Semliki Forest vims vectors for large-scale production of recombinant proteins. Methods... 

Many of these properties would obviously limit applicability of non-continuous cell lines in the industrial-scale production of recombinant proteins. However, such cell types are routinely cultured for research purposes, toxicity testing, etc. 

We prefer large-scale production of recombinant protein by infection in mono-layer culture, since it is easier to achieve synchronous infections in monolayer culture than in suspension culture. 

The use of GHO and NSO cells for the large-scale production of recombinant proteins has been facilitated by the fact that both cell types grow well in single-cell suspension culture with the highest cell densities from an extended batch process reported to be greater than 10 cells mH. This is about five times higher than was seen 20 years ago. Optimized cell lines can achieve secretion of 50 pg per cell per day of recombinant protein. The record on volumetric titer from recombinant mam-... 

All mammalian cells used for the large-scale production of recombinant proteins are considered immortalized , as they can be grown continuously for an indefinite period if correct culture conditions are provided. This is an exceptional characteristic for animal-derived cells, since the tissues and organs of animals are constructed of cells with a defined lifespan. The limited lifespan of cells in animals was detected first by Hayflick [133], and is linked -among other reasons - to a declining telo-merase activity on chromosomes of somatic cells, but not in germ cells. 

Gamier, A., Cote, J., Nadeau, I. et al. (1994) Scale-up of the adenovirus expression system for the production of recombinant protein in human 293S cells. Cytotechnology, 15 (1-3), 145-155. 

In recent years, plants have emerged as a promising new system for the production of recombinant proteins. Plants are widely described as green factories because they provide possible solutions to several of the safety and economic concerns raised by animal systems. The advantages of plants include economical large-scale... 

Recombinant proteins with unique properties can potentially generate new markets and penetrate into existing markets if they can be supplied on a large scale. An ideal system would produce the safest biologically active material at the lowest cost, and would be used in combination with an inexpensive and simple purification process. So far, there have been several examples of the high-yield production of recombinant proteins in transgenic crop plants, mainly in the area of molecular medicines such as antibodies, enzymes and vaccines [45, 48-50]. Modern agricultural practices offer... 

Therapeutic proteins have also been successfully expressed in the urine and seminal fluid of various transgenic animals. Again, issues of sample collection, volume of collected fluid and the appropriateness of these systems render unlikely their industrial-scale adoption. One system that does show industrial promise, however, is the targeted production of recombinant proteins in the egg white of transgenic birds. Targeted production is achieved by choice of an appropriate... 

Historically, HCDC was first established for yeasts to produce single-ceU protein, ethanol, and biomass. Later, dense cultures of other mesophiles producing various types of products were developed, e.g.,by Suzuki et al. [96]. The combination of recombinant DNA technology and large-scale culture processes has enabled human proteins to be produced in a number of hosts, in particular in Escherichia coli [97-100]. Approaches to optimize the production of recombinant proteins are the subject of recent reviews from Winter et al. [101]. 

Mammalian cells are commonly employed for the production of therapeutic and diagnostic proteins, since they are able to correctly synthetize the large and complex structures that the human body requires as medicine [1]. Nowadays, they are employed for the large-scale production of recombinant therapeutic proteins, monoclonal antibodies (MAbs) and viruses used in the preparation of vaccines (e.g. against rabies, hepathytis B, polio, etc) [2]. An overview of some licensed/approved products derived from mammalian cell culture is given in Table 1. 

Large-Scale Cultivation of Host Cells for Production of Recombinant Proteins... 

Figure 4.11. a so liter bench fermenter that can be scaled for production of recombinant proteins. The bench-top scale configuration contains all the control valves and ports necessary to monitor and control cell cultivation while maintaining sterility of the culture. The stainless steal reaction vessel allows easy cleaning and permits heat and pressure sterilization in place by connecting the vessel to a steam supply. (New Brunswick Bioflo-4500, adapted from the manufacturer s literature with permission)... 

Numerous manufacturing systems are presently available for large-scale expression of recombinant proteins. Bacteria, yeast, mammalian cells, and transgenic animals have all been successfully utilized for large-scale production of recombinant biologies. In most cases the recombinant product is a cloned version of a previously purified human or animal protein thus reference standards are available for comparison. The FDA and the European Regulatory Agency have since developed standards for both the production... 

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