Cell-free Expression System for Vaccine Production

Creative Biolabs is an industry leader in vaccine development. We are proficient in all aspects of vaccine development, especially in optimizing the expression system and improving the process. We are convinced that our expertise can help you solve the problems you face in your research.

Cell-free Expression System

The cell-free protein expression system (CFPS) utilizes enzymes and protein factors from cell extracts, exogenous DNA or mRNA as a template, and complements the substrate and energy substances required for translation to achieve in vitro synthesis of the target protein. As early as 1961, Nirenberg and Matthaei have used the cell-free system of E. coli to elucidate the role of the genetic code and its function in protein synthesis. With the deepening of molecular biology and proteomics research and the limitations of traditional cell-based in vivo expression systems in the biopharmaceutical field, cell-free expression systems have once again gained attention and application due to their invulnerability to complex intracellular environment, high tolerance to toxic proteins and low cost of product separation.

In theory, any source of organism can be used to prepare CFPS, but the more mature systems are E. coli extract, wheat germ extract (WGE) and rabbit reticulocyte system (Rabbit reticulocyte extract, RRE). Cell-free expression systems are generally divided into two types: translation systems and couples translation and transcription systems. Both kinds of systems consist of components required for translation like tRNAs, ribosomes, aminoacyl-tRNA synthetases and factors to initiate, elongate and terminate the translation process. In addition, extra supplements such as amino acids, energy sources (ATP, GTP), energy regenerating systems and ions are needed to guarantee efficient translation. As for eukaryotic systems, creatine phosphate and creatine phosphokinase are usually used as energy regenerating system. For prokaryotic systems, phosphoenol pyruvate and pyruvate kinase are commonly added into the system.

Comparison of different cell-free expression systems – Creative Biolabs

Fig. 1 Comparison of in vivo recombinant DNA protein expression with cell-free protein synthesis. (Carlson ED and Gan R, 2012)

There are a variety of cell-free expression systems available categorized into prokaryotic and eukaryotic sources. When choosing an appropriate system for vaccine expressing, researches should take these items into account including the origin of the template RNA or DNA, the expected yield of the product, whether or not demands of post-translational modifications.

System Advantages Disadvantages
Wheat germ 1.Large protein expression
2.No off-target endogenous mammalian proteins
3.Cost-effective preparation of cell extract
4.High yield
1.No mammalian co- and post-translational modifications
2.Premature termination of products
E. coli 1.High yield and production rate
2.Low-cost energy source
3.Tolerant to additives
1.Insoluble expression sometimes
2.No post-translational modifications
3.Different Codon usage from eukaryotes
Insect 1.Large protein expression
2.Glycosylation
1.Non-mammalian
2.Expensive and time-consuming cell cultivation
Rabbit 1.Mammalian system
2.Cap independent translation
1.Sensitive to additives
2.No glycosylation
Human 1.Post-translational modifications
2.Functional protein expression
3.Virus-like particles (VPLs) expression
1.Sensitive to additives
2.Raletively Low yields

Strategies to Improve Expression Level

  • Optimization of the preparation process of cell extract
  • Development of cost-effective energy regenerating system
  • Improvement of stability of the amino acids
  • Enhancement of proper folding and posttranscriptional modifications
  • High-throughput expression of vaccines
  • Individualized customized tumor vaccine

Our Platforms

  • Advanced template design and codon optimization
  • Multiple cell-free expression systems
  • Mature vaccine evaluation system
  • Well-established vaccine developing process

Creative Biolabs is a professional vaccine development expert with extensive experience in vaccine design, immunogenicity evaluation, safety evaluation, and production process optimization. With a variety of mature and comprehensive platforms and technologies for the evaluation and improvement of vaccine expression systems, we can meet the various requirements of our customers.

Reference

  1. Carlson ED. (2012). “Cell-free protein synthesis: applications come of age” Biotechnol Adv. 30(5):1185-94.

Our services are for research use only. We do not provide services directly to individuals.


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