Protein Expression for Custom Antibody Production
Integrated, gene-based protein expression for antigen preparation and custom antibody production
We offer Thermo Scientific Protein Expression and Production Service as a fully-integrated component of our Pierce Custom Antibody Production Services, adding convenience and efficiency to custom antibody development projects.
Our complete set of services for protein expression include gene synthesis (amplification) and subcloning from a variety of sources to create optimized vectors containing appropriate N-terminal or C-terminal tags, transfection or transformation of the recombinant constructs into appropriate cell lines, rigorous testing to identify cells expressing the target protein, large-scale cell culture (production), and final recombinant protein purification using appropriate tag-specific affinity columns.
We use a number of protein expression systems, including bacterial cells (E. coli with PET vector), insect cells (Sf9 cells with pIB/V5-Histadine vector) and mammalian cells (HEK 293 cells with pLOC vector). We can help to design and use the method that is best for the type of antibody production needed. More importantly, we have the world's largest cDNA collection, which provides us immediate access to source coding sequence to construct vectors very quickly and accurately.
- Protein antigen production—use our expression service to generate and prepare protein antigen for a custom antibody production project and acquire the vector to keep
- Optimized vector design—we will suggest the most effective design and expression system to ensure the production of antigen that is effective for antibody production
- Nearly any gene possible—we can amplify and/or subclone from our extensive cDNA library and vector collection, or we can use or obtain sequence from a vector you supply
- Expression systems and affinity tags—choose from bacterial, insect and mammalian protein expression systems and incorporate popular affinity tags
- Protein and vector to keep—any purified protein antigen remaining after completion of the antibody production project is yours to keep, as is the vector constructed
Details and additional information
Protein expression for antigen production
Although protein expression is available in several different species and cell systems, bacterial protein expression is the simplest, fastest and least expensive option for making sufficient protein antigen for antibody production projects.
Therefore, we usually recommend bacterial expression unless one of the following considerations applies:
- The recombinant protein must have specific activity or post-translational modifications (PTM) that only mammalian or insect cell expression can provide
- The intended target and application for the antibody is detection of an E. coli protein
- The customer (for whatever reason) specifically needs a mammalian or insect cell vector as the final deliverable
Our default bacterial protein expression procedure, which typically takes six weeks to complete, is as follows:
- Planning—To avoid any ambiguity, we ask you to supply both protein and nucleotide sequences and to indicate which species you intend for the final antibody to detect. We prefer to use a C-terminal His tag but we can use alternative fusion tags, if desired.
- Construction—In most cases, our library contains the correct cDNA, which we use as the starting material for cloning or subcloning. Otherwise, we can obtain sequence from a plasmid you supply or can synthesize the gene (added cost). For plasmids you supply, we need vector and plasmid sequences, a vector map, and any relevant information.
- Transformation and Selection—We transform recombinant constructs into a high-efficiency expression bacterial strain, induce to overexpress the target protein, select positive colonies, and test to verify expression of the protein.
- Culture and Purification—Finally, we induce two liters of bacterial culture with IPTG, harvest and purify the protein using Ni-NTA (for 6xHis-tagged proteins) or glutathione (for GST-tagged proteins) beads with either native or denatured conditions, as appropriate.
- Delivery—The process typically produces 3 to 5mg of protein, which is sufficient for antibody production projects. If desired, we can supply a small sample of the protein and/or cell stock for you to test before beginning antibody production. Any protein remaining from the antibody production project is delivered to you upon completion.
For Research Use Only. Not for use in diagnostic procedures.