Gibco™ Expi293F™ Cells

Catalog No.	Quantity
A14528	6 x 1 mL
A14527	1 mL

Catalog No. A14528 Supplier Gibco™ Supplier No. A14528

Description

Expi293F human cells are derived from the 293 cell line and are a core component of the Expi293 Expression System.

Expi293F human cells are derived from the 293 cell line and are a core component of the Expi293 Expression System. They are maintained in suspension culture and will grow to high density in Expi293 Expression Medium. Expi293F cells are highly transfectable and generate superior protein yields compared to standard 293 cell lines in transient protein expression.

Expi293F Cells attributes include:

• Suspension culture cells pre-adapted to Expi293 Expression Medium
• Demonstrated higher protein expression levels than other 293 cells adapted to Expi293 Expression Medium
• Exhibit more rapid cell growth and higher culture viabilities than standard suspension-adapted 293 cells
• Consistent transient expression performance of Expi293F cells for over 30 passages beyond thaw
• Also available from a cGMP bank (Cat. No. 100044202)

Frozen cells are supplied in a vial containing 1 mL of cells at 1 x 10⁷ viable cells/mL in Expi293 Expression Medium and 10% DMSO. The cells should be thawed directly into Expi293 Expression Medium. The six-vial multipack eliminates the need to expand and rebank the cells in the lab.

Cells are typically grown in flasks on a shaker platform in a humidified, CO₂ cell culture incubator. However, cultures are scalable from volumes of <1 mL in multiwell plates to >10 L in bioreactors. Protein harvest can be performed at 2–7 days post-transfection, depending on the protein expressed. For expression of recombinant antibodies, harvest is typically at 5–7 days post-transfection. There is no need for culture medium supplementation or batch feeding during expression.

Specifications

• Content And Storage: 1 x 10⁷ cells/mL, store in liquid nitrogen
• Cell Line: Expi293F
• Cell Type: Kidney (Embryonic)
• Culture Type: Suspension Cell Culture
• Form: Cryopreserved
• Species: Human
• Media Recommendation: Expi293™ Expression Medium
• No. of Cells: 1 x 10⁷
• Product Line: Expi293
• Product Type: 293-F Cells
• Quantity: 6 x 1 mL

Frequently Asked Questions (FAQs)

Can I leave my Expi293F culture expressing for more than 7 days?

The growth and expression characteristics of Expi293F cells are such that, by 7 days post-transfection, the culture medium should be close to being spent and maximal protein expression should have already been achieved. Continued incubation will result in a large decrease in cell culture viability.

How many days should I wait before harvesting my protein in the Expi293 Expression System?

The optimal expression time will be different for each protein, but most tested proteins fall within a 3-7 day window. As the system scales well, it is recommended you run a small-scale pilot experiment to determine when to harvest your protein of interest prior to scaling up.

If I let my Expi293F cells grow to a higher density than recommended, will they be okay?

While the Expi293 Expression Medium can support much higher cell densities, we do not recommend growing your Expi293F cultures beyond 5-6 x 10e6 cells/mL, as subsequent transfection and protein expression efficiencies may be reduced. At higher densities, there is also the increased possibility of reaching the point of reduced culture viability. If your seed culture does exceed 5-6 x 10e6 cells/mL, passage them once or twice as detailed in the manual, monitoring them for viability and growth rate. Perform a test transfection using the Protein Expression Control IgG or an expression construct of known yield to determine if cell expression performance has been impacted.

Can I use a different 293 cell line with the Expi293 Expression System?

Other 293 cell lines may be used with the Expi293 Expression System. However, before these cell lines may be used for transfection studies, they must be adapted to serum-free, suspension culture in Expi293 Expression Medium and evaluated for transfection and expression.

Which vector do you recommend using with the Expi293 Expression System?

We recommend using the pcDNA 3.4-TOPO TA vector (Cat. No. A14697). This vector contains the native, full-length CMV promoter and a WPRE (Woodchuck Posttranscriptional Regulatory Element) downstream of the cloning site, both of which contribute to high level gene expression (about 2-3 fold higher expression than with pcDNA3.3 TOPO vector, which in turn provides 2-5 fold higher expression than with a standard pcDNA vector). Of course, the expression level is also protein-dependent.

Is the Expi293 Expression System completely animal-origin free (AOF)?

All the components of the system are animal-origin free except for the Opti-MEM I Reduced Serum Medium that is serum-free but not animal-origin free. Please see the Application Note for using the Expi293 Expression System under animal origin-free conditions:
http://www.thermofisher.com/content/dam/LifeTech/migration/files/proteins-expression-isolation-analysis/pdfs.par.5943.file.dat/expi-293-animal-origin-free-co25751.pdf

I am interested in a mammalian expression system that will allow me to obtain very high yields of my protein. What kind of systems do you offer?

For stable high-yield expression, we offer the Freedom CHO-S and Freedom DG44 kits. For transient high-yield expression, we offer the ExpiCHO Expression System, Expi293 Expression System, FreeStyle 293 Expression System, FreeStyle Max 293 Expression System, and FreeStyle Max CHO Expression System. For high-yield expression of functional membrane proteins in Exp293F cells, we offer the Expi293 MembranePro Expression System (Cat. Nos. A25869, A25870) that combines the scalability and ease of use of Expi293 and the technology of MembranePro to allow an increase of more than 20-fold in membrane protein yield compared to the standard, adherent culture MembranePro Functional Protein Expression System. Please see the Application Note (http://www.thermofisher.com/content/dam/LifeTech/global/life-sciences/ProteinExpressionAnalysis/pdfs/PG1391-PJ6257-CO28704-Expi293Adaption-MembraneProTech_AppNoteGlobal-FHR.pdf) for more details.

What is the biosafety level of Expi293F Cells (Cat. No. A14527, A14528)?

Expi293F Cells is classified biosafety level 2.

As Expi293F Cells (Cat. No. A14527, A14528) are stored in a solution containing 10% DMSO, do they need to be spun down in order to remove the DMSO?

No, they do not need to be spun down to remove the DMSO. As the cells (1 mL) will be transferred to a shaker flask containing 30 mL of pre-warmed expression medium, the DMSO will be diluted so that it won't affect the cells.

Can I create a stable cell line of Expi293F Cells for antibody production using a pcDNA3.4 vector system with G418/Geneticin selection antibiotic?

Please see the following application note "Creation and Scale up of a Stable Cell Line using ExpiCHO Products" (https://assets.thermofisher.com/TFS-Assets/LSG/manuals/MAN0017764_CreatScaleup_StableCellExpiCHO_UB.pdf), that presents our suggested protocol for generation of a stable cell line from ExpiCHO-S cells, which like Expi293F, are grown in suspension.

Do I need to remove Anti-Clumping Agent and Pluronic F-68 from the medium while growing Expi293 cells? How about during transfection of these cells?

You can have Anti-Clumping Agent and Pluronic F-68 in the medium while growing Expi293 cells. You need to remove Anti-Clumping Agent at least 2 passages prior to transfection of Expi293 cells because it interferes with transfection. You can add it back to the medium after transfection. Pluoronic F-68 can be present during transfection as it does not interfere with transfection.

What method do you recommend for quantifying the rabbit Antibody-Expressing Positive Control Vector (Cat. No. A14662)?

The formation of intact IgG molecules may be quantitated using a sandwich ELISA designed to capture and detect rabbit IgG. Besides the rabbit IgG positive control, reagents, and consumables that are included in the kit, you will also need purified rabbit IgG to be used as a standard, F(ab')2 goat anti-rabbit IgG HRP conjugate (Cat. No. A10547), Protein A-coated plates (Cat. No. 15130 for clear plates used in colorimetric detection), TMB colorimetric substrate (Cat. No. 34021), SuperBlock (TBS) Blocking Buffer (Cat. No. 37581), and PBS or TBS buffer for washes. There is an example procedure in our Protein A-coated plates manual (https://assets.thermofisher.com/TFS-Assets/LSG/manuals/MAN0011310_Pierce_ProteinA_G_AG_Coat_96Well_UG.pdf). Please note, our R&D scientists determine titer values from crude cell culture supernatants using a Pall Life Sciences FortéBio Octet instrument equipped with a protein A biosensor.

Safety and Handling

WARNING: Reproductive Harm - www.P65Warnings.ca.gov

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