Production of Recombinant Proteins and Antibodies  


  HEADDr. BÉGUIN Pierre / beguin@pasteur.fr
  MEMBERSDr. BELLALOU Jacques / Dr. BONDET Vincent / Ms. DARTEVELLE Sylvie / Ms. ESNARD Muriel / Mr. FRACHON Emmanuel / Ms. LAURET Marie-Reine
Ms. MARCHAND Françoise / Dr. NATO Faridabano / Ms. NOWAKOWSKI Mireille / Mr. PÊTRES Stéphane


  Annual Report

The Platform is in charge of producing and purifying recombinant proteins and monoclonal antibodies. It comprises three modules :

1. Production of recombinant proteins in microbial systems.

The module performs bacterial cultures in shake flasks, in classical 2- and 15-liter fermentors and in a micro-fermentor battery that was designed and developed in-house for performing parallel cultures under defined conditions (Figure 1). The device features an optical system enabling on-line monitoring of the optical density of the cultures. A license has been granted to the company Fogale nanotech, who will soon market a commercial version of the battery.

In addition to performing to cultures under standardized conditions, a major share of the activity of the module consists in optimizing the production of "difficult" proteins that are synthesized in low amounts or in insoluble form.

2. Production of recombinant proteins in insect cells.

The Platform also offers the opportunity to produce recombinant proteins in baculovirus-infected cells of Spodoptera frugiperda or in stable transformed lines of Drosophila melanogaster Schneider cells. For the baculovirus system, the module performs the construction of the recombinant virus starting from a transfer vector and the production of high-titer stocks. This kind of service faces increasing demand, particularly for proteins from higher eukaryotes, such as proteins from parasites and viruses, which cannot be conveniently produced in an active form in microbial systems.

3. Production of monoclonal antibodies

The module uses the classical technique of Kohler and Milstein to produce mouse monoclonal antibodies against antigens provided by the users. Antibodies are characterized with respect to their Ig subtype and their dissociation constant. Epitope mapping can be performed if the adequate antigens are provided.

Currently, the module manages about a dozen projects at various stages of completion. Some 6-7 projects involve the production of monoclonal antibodies for research. One project seeks to obtain neutralizing antibodies against Clostridium botulinum toxins. The remaining projects concern the development of diagnostic tests using dipsticks based on immunochromatography for the detection of pathogen-specific antigens in biological fluids. These include diagnoses for bloody diarrheas, meningitis, plague, and Plasmodium falciparum.

Running procedures

Short-term projects (<1-2 weeks) are treated in line; longer projects are submitted to a steering committee for approval. Applicants are expected to shoulder the running expenses incurred by the project.

Keywords: bioreactors, recombinant protein, monoclonal antibodies, dipstick diagnosis, baculovirus

Ptprotrec.jpg

General view of the micro-fermentor battery.



  Publications

Maillard P, Lavergne JP, Siberil S, Faure G, Roohvand F, Petres S, Teillaud JL, Budkowska A. 2004. Fcγ receptor-like activity of hepatitis C virus core protein..J Biol Chem. 279, 2430-2437.

Frachon E, Bondet V, Munier-Lehmann H, and Bellalou, J (2006). Multiple micro-fermentors: a versatile tool for automated parallel cultures of microorganisms producing recombinant proteins and for optimization of cultivation protocols. Appl. Environ. Microbiol. 72, 5225-5231.

Chanteau S, Dartevelle S, Mahamane AE, Djibo S, Boisier P, Nato F. (2006). New rapid diagnostic tests for Neisseria miningitidisi serogroups A, W135, C, and Y. PLoS Med. 3, e337

Phalipon ., Costachel C, 2, Grandjean C, Thuizat A, Guerreiro C, Tanguy M, Nato F, Vulliez-Le Normand B, Bélot F, Wright K, Marcel-Peyre V, 2, Sansonetti PJ, Mulard LA. 2006. Characterization of functional oligosaccharide mimics of the Shigella flexneri serotype 2a O-antigen: implications for the development of a chemically defined glycoconjugate vaccine. J. Immunol. 176, 1686-1694.

Nato F, Phalipon A, Nguyen TL, Diep TT, Sansonetti P, Germani Y. 2009. Dipstick for rapid diagnosis of Shigella flexneri 2a in stool. PLoS One 2, e361.





Activity Reports 2007 - Institut Pasteur
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