Recombinant Protein Expression in Mammalian Cells
Mammalian expression system are the primary option for the production of recombinant proteins and antibodies in pharmaceutical research. Mammalian cells can produce recombinant proteins with proper post-translational modifications and folding, which are important for biological activity. In addition, mammalian expression system doesn’t have endotoxin problem in E.coli expression system. Commonly used cell lines include human 293 cells and CHO.
Bon Opus has decades of experience in mammalian cell expression. Our in-house protein experts will help you explore different options and work closely with you each step of the way.
Feature Package: Pilot-scale Mammalian Protein Expression ($950)
Include gene synthesis, cloning, transfection-grade DNA preparation
Transfection and expression in 20ml cell culture
One-step purification with tag (His or Fc)
6-8 weeks delivery time
1. Gene synthesis
2. Cloning & Transfection
Signal peptide is critical for secretion in mammalian cells. Based on the protein of interest, we will help determine the best signal peptide to use. The options include common signal peptide such as IL-2, CD33 leader sequence, as well as our in-house developed signal peptide sequences.
The addition and position of epitope tags will be decided by the application of the protein. Commonly used tags include His, FLAG, and Fc.
Gene Synthesis & Cloning
Once a protein of interest is identified, the AA will be subjected to codon optimization to increase translational efficiency. The optimized gene will be synthesized and sub-cloned into our in-house developed expression vector. The entire process can be completed within 2 weeks.
Once an expression vector is made, a trial transfection will be conducted to determine the rough yield of the protein. The purified samples from the pilot can also be used to test its bioactivity in POC experiment.
Scale-up transfection will be performed according to the predicted yield and quantity need. It normally takes 2-3 weeks to complete. Based on the epitope tags and purity demand, affinity purification (protein A and his) along with other purification methods (IEX, gel-filtration, HIC, etc.) will be carried out to purify the target protein.
After all procedures are completed, the proteins will be subjected to a round of final QC, which can include SDS-PAGE (reducing and non-reducing), concentration measurement (BCA, Bradford or A280), western blot, SEC-HPLC, endotoxin removal lowest to 0.1EU/mg.