Custom Expression Systems
MDS's custom expression group provides a fully integrated and documented service from project design to execution for research, development, and scale up. These services will take you from optimization through expression of a cloned sequence to purification and characterization of the expressed protein. MDS offers a choice of several systems and various services.
• Mammalian expression systems
• Prokaryotic expression systems
• Transfection and purification
• Generation of high-titer stocks
• Expression services
• Large-scale expression and processing
Mammalian cell lines offer several advantages for expression of heterologous proteins. Since transcription, translation, and post-translational modification processes are conserved among higher eukaryotes, eukaryotic proteins produced in mammalian cells will be functional and well suited for a variety of structure-function assays and analysis of the physiological effects of the protein on cell function. These services include transfection of vector DNA into mammalian cells for stable or transient expression of gene products. A variety of transfection methods and assay systems are available.
Custom expression in insect cells and lytic baculoviruses provide a proven method for high-level expression of full-length mammalian proteins, providing researchers with the means to generate milligram quantities of cloned proteins that are ideal for functional studies and structural analyses.. Many post-translational modifications similar to those in mammalian cells are made in insect cells, and proteins unable to be expressed in E. coli have been successfully expressed using the baculovirus system . MDS's services include cloning of the gene of interest into a viral vector such as vaccinia or bacculovirus and small or large-scale production of infected cultures for isolation of the expressed protein
Prokaryotic Expression Systems
MDS has expertise in using a variety of systems for expression of recombinant proteins in E. coli . Each system offers expression vectors with unique characteristics that help you tailor your experiments to fit the protein of interest.
- Transformation with Client supplied vector
- Growth and induction; screening for colonal variation of intracellular or secreted product
- Cell bank preparation
- Shake flask production with SDS PAGE and Western blot analysis
- Production in 5 and 10 L fermenters
Generation of high-titer stocks
A purified recombinant (provided by client) will be utilized to infect the cells and generate one to five liters of high-titer stock. The titer will be determined by plaque assay. The outline of procedures is summarized below.
• Maintenance of infection for the optimal time of expression as specified by the client (predicted by previous analysis or time course study)
• Harvesting of cells and supernatant at optimal time point (specified by client). Submission of entire supernatant and cell pellet to the client (as specified in advance).
The outline of a time-course study is summarized below. Contact MDS for custom analysis
• Infecting the cells at log phase with inoculum of previously generated and titered high-titer stock.
• Maintenance of time course infections with time point samples (cells and supernatant) taken every 24 hours for 1-5 days.
• Time point samples, including cells and supernatant from recombinant infections, will be sent to the researcher for analysis and determination of the optimal time for expression and harvesting of recombinant proteins.
These services include growing one cell line (specified by the client) to log phase and infecting the cells with previously generated high titer stock. Fermentation optimization to increase expression is available in bacteria, yeast, insect, and mammalian cell lines.
Large-scale expression and processing
MDS offers large-scale expression and processing of recombinant proteins.
MDS's Technical Services Division offers many additional services to save you valuable time and resources. Additional services include purification of expressed protein (yields may be improved through addition of affinity tags) and analysis of expressed protein by SDS-PAGE and Western blot (utilizing your specific antibody). Filling, capping, and labeling services are also available.
Pricing is based on the complexity of each project, the type of selected cell, and the services rendered. MDS's custom protein expression, purification, and characterization services can be tailored to suit your needs.