Has the DG44 cell line been used for the large scale production of any FDA-approved biotherapeutic proteins?
70% of all therapeutics produced in mammalian cells are expressed in CHO cells. CHO cells are preferred because of human-like glycosylation and no propagation of human-like viruses. Most therapeutic IgGs are produced using the CHO platform and a number of FDA-approved biotherapeutics have been produced in DG44 cells.
Why are the defrosted CHO DG44 Cells (cGMP banked) transferred directly into pre-warmed media instead of being centrifuged for recovery when using the CHO DG44 Cells (cGMP banked) and Media Kit (Cat. No. A1100001)?
The defrosted contents of the vial of Gibco CHO DG44 Cells are transferred directly to 29 mL of pre-warmed media and then incubated because the cells are extremely fragile upon recovery from cryopreservation. Centrifuging the cells at this stage could cause significant damage, reducing their viability and recovery efficiency. To help ensure optimal recovery and viability, the cells are directly transferred to pre-warmed media and incubated under optimal conditions.
This approach is also why we do not mention centrifuge resuspension in the subculture section. We have found that the shear forces during centrifugation are more harmful to the cells than retaining a small amount of old media or DMSO. The volume of media in the flask has been found to sufficiently dilute the DMSO such that it does not impact recovery or cell growth rather than using a centrifuge.
How can I learn more about commercial licensing of Freedom kits, ExpiCHO-S Cells (cGMP banked), CHO-S Cells (cGMP banked), and CHO DG44 Cells (cGMP banked)?
To inquire about a Commercial Production or Service License, please email: outlicensing@thermofisher.com.
What is the term of the license agreement for ExpiCHO-S Cells (cGMP banked), CHO-S Cells (cGMP banked), and CHO DG44 Cells (cGMP banked)?
Licensing rights are granted in perpetuity for the Commercial Production License. The term of the Commercial Service License is negotiated between the parties.
Is there a pull-through royalty on final products or an annual maintenance fee for using ExpiCHO-S Cells (cGMP banked), CHO-S Cells (cGMP banked), and CHO DG44 Cells (cGMP banked)??
Only a one-time license fee is required for a Commercial Production License, and it covers use during the clinical trial process and commercialization of an unlimited number of molecules.
Are separate licenses required for ExpiCHO-S Cells (cGMP banked), CHO-S Cells (cGMP banked), and CHO DG44 Cells (cGMP banked)?
Yes. Each cell line requires a separate Commercial Production or Service License.
When is a license required for ExpiCHO-S Cells (cGMP banked), CHO-S Cells (cGMP banked), and CHO DG44 Cells (cGMP banked)?
A Limited Use Label License, which includes research rights, is available in the product manual and applies when the cells are purchased. An additional Commercial Production or Service License is required in the following circumstances:
- A Commercial Production License is required prior to the clinical use in humans of expressed proteins, and/or when an IND submission is being prepared for first in human clinical studies.
- A Commercial Service Provider License is required by service providers prior to use of cells for any client projects.
Is there information on ExpiCHO-S Cells (cGMP banked), CHO-S Cells (cGMP banked), and CHO DG44 Cells (cGMP banked) to support regulatory filings?
Yes. After execution and payment for a Commercial Production License, a Cell Line Documentation Package (>200 pages, and specific to the lot of cells purchased), is provided which includes the entire lineage history of the cells starting at receipt of the initial vial at Thermo Fisher Scientific through delivery to the purchaser, as well as all testing reports.
What fields of use are allowed for ExpiCHO-S Cells (cGMP banked), CHO-S Cells (cGMP banked), and CHO DG44 Cells (cGMP banked)?
Many fields of use are available, including development and production of molecules in the area of human biotherapeutics, human vaccines, veterinary vaccines, human diagnostics, or animal diagnostics, and production of proteins for sale into research markets. For more details about fields of use, please contact us at outlicensing@thermofisher.com.
What Freedom kit cell types are available for commercial licensing?
ExpiCHO-S Cells (cGMP banked), CHO-S Cells (cGMP banked), and CHO DG44 Cells (cGMP banked) can be licensed for commercial applications.
Do you offer technical support for the Freedom DG44 Kit?
Yes. Contact our field-based Process Science Fellows (PSFs) at pd.direct@lifetech.com for technical support.
How can I learn more about commercial licensing of CHO DG44 Cells (cGMP banked)?
To inquire about a commercial license, please email: pd.direct@lifetech.com. To read the complete list of FAQs related to licensing, go here (https://www.thermofisher.com/us/en/home/life-science/bioproduction/bp-misc/gibco-freedom/freedom-cho-s-cells-and-freedom-dg44-cells-licensing.html).
Is there a pull-through royalty on final products or an annual maintenance fee when using the Freedom DG44 Kit??
No. Only a one-time license fee is required, and it covers the commercialization of an unlimited number of molecules using the CHO DG44 Cells (cGMP banked).
When is a license required for the Freedom DG44 Kit?
A commercial use license is required prior to the sale of expressed proteins, or when an IND submission is being prepared for first in human clinical studies.
What are some considerations for successful scale-up of CHO DG44 Cells (cGMP banked)?
IgG-expressing CHO DG44 clones readily scale-up from shake flask to bioreactor vessels while maintaining growth and titer performance.
What are some considerations for obtaining a high number of stable clones using the Freedom DG44 Kit?
When using limiting dilution cloning, careful evaluation of monoclonality of a clone is required before assessing stability of expression in order to avoid heterogeneous clones that can drift into separate populations and appear unstable.
How many transfections are typically required to get high-expressing clones from the Freedom DG44 Kit?
Only two transfections are recommended.
What transfection methods can be used with the Freedom DG44 Kit?
Both FreeStyle MAX Reagent (included in the kit) and the Neon Transfection System provide stable pools with similar final titers
What cloning methods can be used with the Freedom DG44 Kit?
Good results can be obtaining using either limiting dilution cloning or ClonePix FL. For advice regarding ClonePix cloning, see recommendations in the Freedom DG44 Kit manual or request a consultation at pd.direct@lifetech.com
What selection system is used in the Freedom DG44 Kit?
In the Freedom DG44 Kit, both Geneticin and DHFR selection are used, either sequentially or simultaneously.
Where are the Freedom DG44 Kit vector maps located?
Detailed information about the pcDNA 3.3 TOPO TA vector and pOptiVEC TOPO TA vector can be found in the Freedom DG44 Kit manual.
Does the Freedom DG44 Kit contain documentation for the CHO DG44 Cells (cGMP banked)?
A certificate of analysis (COA) demonstrating suitability for research use is available to everyone who purchases the CHO DG44 Cells (cGMP banked) and Media Kit or the Freedom DG44 Kit. A full documentation package, including the cell line's lineage, history, and cGMP testing results, is offered in conjunction with a commercial use license
What is included in the Freedom DG44 Kit?
All major components needed for the development of stable clones are included in the Freedom DG44 Kit: CHO DG44 Cells (cGMP banked), vectors for cloning one or two genes (pcDNA 3.3 TOPO TA Vector Kit and pOptiVEC TOPO TA Vector Kit), transfection regent ( FreeStyle MAX Reagent), competent cells (One Shot TOP10 Chemically Competent E. coli), serum-free cell culture media (CD DG44 Medium, CD OptiCHO Medium, and OptiPRO SFM), and other reagents such as Pluronic and Gentamicin. A complete list of components, including additional materials required but not included, can be found in the Freedom DG44 Kit manual.
Can CHO DG44 Cells (cGMP banked) be used for the production of commercial products?
Yes. CHO DG44 Cells (cGMP banked) are intended (with the purchase of a commercial license) for the development of biologics. They have been produced in a cGMP animal origin-free environment, and have been scaled up to produce products for clinical testing.
How successful is the Freedom DG44 Kit in creating stable clones?
The stability of expression was assessed for over 60 generations with the observation that most clones maintain their level of expression after 60 generations. Expression levels were considered stable if the titer at the end of the stability assessment (at generation 60) was at least 70% of the titer observed at the beginning of the stability study.
What level of product quality is seen with the Freedom DG44 Kit?
Product quality depends on multiple development aspects, such as the product itself, the selected clone, and the upstream and downstream process. CHO DG44 Cells (cGMP banked) offer all the typical CHO cell line benefits with respect to low protease levels and preferred glycan pattern.
How long does it take to obtain a lead clone with the Freedom DG44 Kit?
It is possible to go from transfection to lead clone in less than 6 months with the Freedom DG44 Kit.
What titers can be achieved with the Freedom DG44 Kit?
IgG titers of ≥0.5 g/L in simple fed batch and ≥1 g/L in fed batch with complex feeds have been obtained using the Freedom DG44 Kit. Titers of 20-100 mg/L with blood factors and other hard to express proteins (HTEP) have also been achieved with this kit.
