Fast, reliable protein expression and production from mammalian or insect cells

Recombinant protein expression can be used across diverse areas of life science research─from the analysis of cellular proteins to high-yield protein production for drug development. For those interested in outsourcing protein expression and purification steps, our GeneArt™ protein purification services offer a rapid and dependable way to obtain correctly folded native protein from transiently transfected mammalian or insect cells. We use optimized conditions throughout the workflow, enabling us to routinely and efficiently obtain high yields of purified proteins.

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General step-by-step guide for GeneArt protein purification services

Protein sequence and project requirements

The basic requirements for initiating your project are: 

  • An electronic file of the protein sequence of interest 
  • Detailed project specifications

DNA optimization and generation of expression construct 

To help ensure optimal gene expression and maximal design flexibility, we recommend starting every protein expression project with an expression-optimized gene sequence and optimized expression vector. 

  • DNA sequence optimization by GeneOptimizer™ software, which preserves the protein amino acid sequence, is proven to result in higher and more reliable expression levels for many proteins (Fath et al., 2011).
  • To help get maximal expression, we recommend using a vector optimized for the expression system. You can send us your own expression vector, or we can assist with selecting one of our expression-optimized vectors.

Learn more about:

  Advanced expression vectors

Plasmid preparation

To provide transfection-ready constructs, our experienced scientists use proven protocols and quality-controlled systems and reagents to prepare optimal purified plasmid DNA (pDNA). 

Learn more about:

 

Transfection and expression

The combination of an optimized expression plasmid with our advanced mammalian expression systems that are adapted for high-yield recombinant protein expression can make a difference for your results. 

  • Optimized transfection and cell culture protocols minimize the risk of process failures and delays. 
  • Depending on the expression system and expressed protein, culture time can vary between 2 and 14 days and can be customized if needed. 
  • Culture supernatant or cells are harvested and rapidly and efficiently advanced to protein purification. 

Learn more about optimized expression systems and transfection conditions: 

Purification

We typically use state-of-the-art purification protocols, but we can also use a customized protocol sent to us. 

  • The first round of purification is done routinely via affinity chromatography (e.g., using an Fc or His tag). 
  • A second round of purification (e.g., size exclusion chromatography (SEC) via HPLC) can be added to increase purity, if necessary.  

Service features:

  • Affinity chromatography (several tags available)
  • Additional polishing steps, as needed, for highly purified proteins

Documentation and delivery

As standard procedure, several basic quality control (QC) steps are performed and documented in detail for all protein expression and purification projects. This includes analyzing samples by SDS-PAGE and western blot. Based on your specific project requirements we can offer additional QC methods, including endotoxin testing and analytical SEC via HPLC. We can customize all documentation as necessary. Upon successfully passing all QC checks, purified proteins are released for shipment. 

Deliverables:

  • Purified protein and expression vector used for transfection
  • Detailed documentation including data from SDS-PAGE and western blot analyses

 

   

How to get your project started

  Download and fill out the Protein Production Request Form, then email it to the GeneArt Protein Services team at geneartprotein@thermofisher.com. We will respond with a price and production time quote. (Pricing depends on project details and will be provided after inquiry.)

Why GeneArt protein purification services?

 

All in-house production: Each step from oligo synthesis to the purified protein Confidentiality and no need to coordinate different vendors
Short processing time: Gene to protein, starting from 30 business days

Reliable and advanced expression systems: FreeStyle™ 293, FreeStyle™ CHO, Expi293™, ExpiCHO™, Bac-to-Bac™ cell lines. All reagents commercially available
Superior gene expression optimization: Proprietary GeneOptimizer™ software

Range of scales: From 30 mL to 25 L for research scale
Customized service: Ready-to-use customer purification protocols, expandable capabilities

Dedicated and experienced project management team: Close communication in all project phases, experience with large projects:
Active MSA with large pharma
Secure E-business solutions: unique portal

Service Description Deliverables
Gene to protein
Specified culture volume
Protein expression and purification from customer-specified culture volume of transiently transfected HEK293, CHO, or insect cells
  • All protein purified from specified culture volume (alternatively, culture supernatant or cells)
  • Documentation including data from SDS-PAGE and western blot
  • Expression plasmid
Gene to protein
Specified protein amount
  • Protein expression and purification of customer-specified guaranteed amount of protein using transiently transfected HEK293, CHO, or insect cells
  • Pilot expression service is mandatory
  • Purified protein in amount specified by customer
  • Documentation including data from SDS-PAGE and western blot
  • Expression plasmid
Gene to protein
Pilot
Feasibility study for determination of production yield from transiently transfected HEK293, CHO, or insect cells
  • Price quote for production of a customer-specified amount of protein
  • Documentation including data from SDS-PAGE and western blot
  • All resulting protein from pilot study
  • Expression plasmid

Please inquire with our support team at geneartprotein@thermofisher.com for your project setup and quote. 

Service project phase or characteristic Standard specifications
Mammalian expression systems
  • Invitrogen FreeStyle CHO-S cells
  • Invitrogen FreeStyle 293-F cells
  • Gibco Expi293 F cells
  • Gibco ExpiCHO-S cells
Insect expression system
  • Invitrogen™ Bac-to-Bac™ expression system
Expression scales
  • 30 mL to 25 L
  • Shaker and WAVE™ bag–based cell culture
Purification methods
  • Affinity chromatography
  • Size exclusion chromatography (SEC)
  • Ion exchange chromatography (IEX)
Affinity tags
  • His-Tag
  • Fc-Tag (Protein A/G)
  • Strep-Tag
  • GST-Tag
  • Other, upon request
Additional options
  • 2nd column purification
  • Tag cleavage
  • Analytical SEC (HPLC)
  • Customized buffer
  • Endotoxin determination
  • Aliquot service
  • Concentration adjustment
  • Titer determination in culture supernatant (antibodies)
  • Other, upon request

Please inquire with our support team at geneartprotein@thermofisher.com for your project setup and quote.

Quality control specifications Documentation
SDS-PAGE, visualization of protein by Coomassie blue staining Included in standard QC
Spectrophotometric determination of protein concentration Included in standard QC
Western blot with tag-specific antibody Included in standard QC
Capillary gel electrophoresis, detection by UV absorbance and/or fluorescence Upon request
Analytical SEC (HPLC) Upon request
LAL assay for endotoxin determination Upon request

Case study: Improvement of antibody expression after sequence optimization with GeneOptimizer software

In this case study we compared the expression levels of two antibodies derived from optimized sequences, relative to their wild type counterparts. The open reading frames (ORFs) of the two antibody-encoding genes were optimized for human expression using the GeneOptimizer algorithm. Gene synthesis, subcloning into the Invitrogen™ pcDNA™ 3.3 vector, and plasmid preparation were performed in-house. The four resulting constructs (for each of the two antibodies, one wild type and one sequence-optimized) were expressed in duplicate in Invitrogen™ FreeStyle™ 293 cells at 30 mL scale. The expression levels were determined using the ProtA BLItz® system (ForteBio, PALL Corp.).

Analysis of expression after normalization to wild type levels clearly showed a strong increase for the sequence-optimized antibodies (Figure 1). This means that high levels of purified protein can be obtained rapidly and efficiently at lower overall project costs.

Expression of wild type vs. sequence-optimized antibodies
Expression of wild type vs. sequence-optimized antibodies

Figure 1. Expression of wild type vs. sequence-optimized antibodies. Mean protein expression was normalized to the mean expression of the wild type sequences.

 

Customized service

If you already have an optimized expression construct and/or expression protocols available, we can also use these to purify your protein in our facilities. Please contact geneartprotein@thermofisher.com for further details.

For Research Use Only. Not for use in diagnostic procedures.