Recombination cell culture

Mammalian cells Cultured human lymphocytes Mitotic recombination Aberrant metaphases No data - L Vova 1984... 

Significantly, therefore, recombinant DNA and hy-bridoma techniques are capable of providing molecules of a well-defined chemical composition and producing them in cell culture media that can be carefully controlled. In short, the critical advance these techniques offer is that supply and purity no longer impede the development and clinical usefulness of therapeutic protein molecules. 

In order to reduce the time required to confirm the accumulation of a given recombinant protein, we have developed a cell culture system in which transgenic alfalfa callus material produced at the proliferation step of Agrobacterium-based transformation is used to initiate cell cultures. These cell suspensions can be subcultured to sustain batch production of modest protein amounts. The protein blot shown in Fig. 1.2 demonstrates our ability to detect a recombinant protein in total... 

When sufficiently high levels of expression and protein accumulation are achieved, efficient downstream processing protocols must be developed to insure product quality and the economic feasibility of production. As the demand for safe, recombinant pharmaceutical proteins continues to expand, the market potential of plant-produced recombinant proteins is considerable. Molecular farming can produce recombinant proteins at a lower cost than traditional expression systems based on microbial or animal cell culture, and without the risk of contamination with human pathogens. 

The third time constraint depends on whether the product can be extracted from seeds or fruits. This uncouples protein expression and purification. Large batches of seeds containing the recombinant protein can be produced and stored at low costs. Provided the protein remains stable in the stored seeds, purification can be carried out on demand or shifted according to free capacities. The advantage of one large harvest, with seeds mixed to uniformity, is that this allows production on demand. In contrast, mammalian cell culture is prone to minor batch-to-batch variations in... 

Stable expression of the recombinant protein is another pre-requisite. For production in CHO cell culture, narrow specifications of expression level are requested by the FDA. Consequently, plant-derived proteins need to be produced according to a defined scheme and within defined specifications in order to generate material suitable for clinical trials. Therefore, the expression level must be maintained within a defined range. For example, Prodigene Inc. assessed whether the E. coli Lt-B protein was uniformly distributed throughout the defatted maize germ by analyzing samples... 

Expression of recombinant proteins in animal cell culture systems... 

In addition to recombinant biopharmaceuticals, animal cell culture is used to produce various other biologically based pharmaceuticals. Chief amongst these are a variety of vaccines and hy-bridoma cell-produced monoclonal antibodies (Chapter 13). Earlier interferon preparations were also produced in culture by a particular lymphoblastoid cell line (the Namalwa cell line), which was found to synthesize high levels of several IFN-a s naturally (Chapter 8). 

Although capable of glycosylating heterologous human proteins, the glycosylation pattern usually varies from the pattern observed on the native glycoprotein (when isolated from its natural source, or when expressed in recombinant animal cell culture systems). 

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