Efficient Small-Scale Production of Proteins - - BioPharm International


Efficient Small-Scale Production of Proteins

BioPharm International

Efficient gene transfer into cells relevant for protein production purposes has been a major bottleneck. Viral gene transfer systems usually have the advantage of high transduction efficiencies compared to non-viral methods. However, these methods suffer from several limitations such as the time-consuming and laborious production of vectors, elevated laboratory costs due to the high level of safety requirements, limitation of insert size, and possible immunogenic reaction in clinical trials.4

Figure 2. Efficient Gene Transfer Using Nucleofection.
Non-viral gene transfer methods include calcium phosphate, lipofection reagents, electroporation, and ballistic gene transfer.5 Whereas electroporation and ballistic techniques usually lead to high cell mortality, calcium phosphate and lipofection often result in low transfection efficiencies especially in suspension cells. The electroporation-based Nucleofector technology is a valid alternative and has been proven to be efficient even in suspension cells relevant for protein production (Figure 2).6

Stable protein production in mammalian cells

Cells used for the production of therapeutic proteins must comply with various requirements to ensure approval for the protein as a drug. One of those requirements is to use a thoroughly defined clone. It must be guaranteed that the cells used for protein production are derived from a single clone with a specifically-defined integration site. The generation of a stable clone often requires six months or more due to selection procedures and adaptation to serum-free conditions (Figure 3A). A variety of systems for selecting transfected cells exists, including resistance to antibiotics such as neomycin, hygromycin and puromycin, dihydrofolate reductase (DHFR) and glutamine synthetase (GS) systems.

Figure 3. A. The generation of a stable clone requires six months or more due to selection procedures and adaptation to serum-free conditions (SFM). B. If serum-free adapted suspension cells can be transfected, much time can be saved in the later process.
The aim of selection is to identify high producing clones, but this is a tedious and labor-intensive exercise. Several methods for isolating clones are used, the most popular one being cloning by limiting dilution using multiwell plates. Once a stable clone is selected, it must be adapted to serum-free suspension culture so that it can be used in efficient large-scale production. This adaptation process is again time consuming and bears the risk that the clone will lose its desired high producing properties. By applying novel transfection technologies such as the Nucleofector technology, suspension cells can be transfected directly in a serum-free environment. This saves significant time and reduces the risk on the way to find the right clone (Figure 3B).

Transient Protein Production in Mammalian Cells

blog comments powered by Disqus



Compounding Pharmacy Issues Recall, But Challenges FDA Decision
July 22, 2014
AbbVie's Acquisition of Shire Could Save $8 Billion in Taxes
July 21, 2014
AbbVie to Acquire Shire for $54.7 Billion
July 18, 2014
AstraZeneca Reveals Design for New Global R&D Center and Corporate Headquarters
July 18, 2014
Particulate Matter Prompts Baxter's Recall of IV Solutions
July 17, 2014
Author Guidelines
Source: BioPharm International,
Click here