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Gibco™ Expi293™ Inducible GnTI- Expression System Kit

Product Code. 16396395
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Product Code. 16396395 Supplier Gibco™ Supplier No. A39252

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Description

Expi293 Inducible GnTI- Expression System Kit is part of the Expi293 platform for rapid, high-yield transient protein expression from mammalian 293 cells

  • System kit is centered on engineered Expi293F cells that do not have N-acetylglucosaminyltransferase I (GnTI) activity, therefore lacking complex N-glycans, and stably express the tetracycline repressor (TetR) protein, thus enabling tetracycline-regulated protein expression
  • Expi293F Inducible GnTI- cell line was established by the selective knockout of the GnTI gene and the stable integration of the regulatory plasmid pcDNA6/TR, which encodes the TetR gene under the control of the human CMV promoter, in parental Expi293F cells
  • This cell line is a powerful tool for tetracycline-inducible high-yield expression of homogeneously glycosylated recombinant proteins.

Features:

  • Homogeneous N-glycosylation of expressed proteins
  • Tetracycline-inducible protein expression with extremely low basal expression levels
  • Equivalent protein yields to parental Expi293F cells—up to 1 g/L
  • pcDNA5/TO expression vector for tetracycline-regulated expression of the gene of interest
  • Suspension, high-density culture of Expi293F Inducible GnTI- cells in Expi293 Expression Medium
  • High-efficiency transient transfection using ExpiFectamine 293 reagent in combination with specialized transfection enhancers

Components included:

  • 1 vial of frozen Expi293F Inducible GnTI- cells
  • 1 L of Expi293 Expression Medium
  • 1 ExpiFectamine 293 transfection kit sufficient to transfect 1 L of culture,
  • 100 mL Opti-Plex Complexation Buffer
  • 1  PNGase F Glycan Cleavage kit
  • 1 pcDNA5/TO Mammalian Expression Vector kit
  • 500 mg tetracycline hydrochloride
  • pRABBIT IgG IRES-EmGFP positive control vector

All components of the Expi293 Inducible GnTI- Expression System are also available for individual purchase

TRUSTED_SUSTAINABILITY

Specifications

Delivery Type Transfection
Quantity 1 kit
Content And Storage • Expi293F Inducible GnTI- Cells (1x107 cells/vial), store in vapor-phase liquid nitrogen
• 1 L Expi293 Expression Medium, store at 2–8°C (protect from light)
• ExpiFectamine 293 Transfection Kit for 1 L of culture, store at 2–8°C
• PNGase F Glycan Cleavage Kit, store at -20C
• 100 mL Opti-Plex Complexation Buffer, store at 2–8°C (protect from light)
• 20 μg pcDNA5/TO Mammalian Expression Vector, store at 2–8°C or long term storage at -20°C.
• 500 mg Tetracycline Hydrochloride, store at -20°C
• pRABBIT IgG IRES-EmGFP Positive Control Vector, store at 2–8°C or long term storage at -20°C
Shipping Condition Approved for shipment at Room Temperature or on Wet or Dry Ice
Cell Line Expi293F™
Product Line Expi293
For Use With (Application) Protein Production
Promoter CMV/TO

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.
What is the difference between the Expi293F Inducible Cell Line and the T-REx -293 Cell Line?

The Expi293F Inducible Cell Line was created using the same technology as the T-REx -293 Cell Line. Both cell lines have been created by stable transfection with the pcDNA 6/TR vector; thus, they ubiquitously express the tetracycline repressor (TetR) protein. The parental cell line of Expi293F Inducible cells is Expi293F Cells (Cat. No. A14527). The key differences between these 2 cell lines are:
- Expi293F Inducible cells are grown in suspension and are adapted to high-density culture (> 15x10E6 cells/mL).
- Expi293F Inducible cells grow in Expi293 Expression System and perform equivalently to parental Expi293F cells, thus making them ideal for high-yield, inducible protein expression.
What method do you recommend for quantifying rabbit IgG expression from pRABBIT IgG IRES-EmGFP Positive Control Vector, Cat. No. A39243?

The formation of intact IgG molecules may be quantified using a sandwich ELISA designed to capture and detect rabbit IgG. Besides the rabbit IgG positive control vector, 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 (H+L) Cross-Adsorbed Secondary Antibody, HRP (Cat. No. A10547), Protein A Coated Plates, Clear, 96-Well (Cat. No. 15130), TMB Substrate Kit (Cat. No. 34021), SuperBlock (TBS) Blocking Buffer (Cat. No. 37535), and PBS or TBS buffer for washes. There is an example procedure in our Protein A Coated Plates manual (https://tools.thermofisher.com/content/sfs/manuals/MAN0011310_Thermo Scientific_ProteinA_G_AG_Coat_96Well_UG.pdf). Please note that 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.
Do you offer a GFP-expressing mammalian expression vector that I can use as a control to monitor my transfection and expression with the Expi systems?

We offer pRABBIT IgG IRES-EmGFP Positive Control Vector, Cat. No. A39243, which you can use to monitor your transfection and expression.
Can I use doxycycline instead of tetracycline as an inducer in the Expi293 Inducible Expression System?

Doxycycline may be used as an alternative inducing agent in the Expi293 Inducible System. It is similar to tetracycline in its mechanism of action. Doxycycline has been shown to have a longer half-life than tetracycline (48 hr vs. 24 hr, respectively). If the use of doxycycline is desired with the Expi293 Inducible System, we recommend performing a dose-response curve to determine the optimal concentration for induction.

How much tetracycline do I need to induce expression in Expi293F Inducible or Expi293F Inducible GnTI- cells?

We recommend starting with a tetracycline concentration of 1 µg/mL to induce the expression of your protein of interest. However, you can modulate the level of expression by treating cells with different amounts of tetracycline. We recommend performing a dose-response curve (0-1 µg/mL; 2-fold dilution) to determine the optimal concentration of tetracycline required to induce your protein of interest.
What vector do you recommend using with the Expi293 Inducible Expression System?

We recommend using the pcDNA 5/TO Mammalian Expression Vector (Cat. No. 430096). The pcDNA 5/TO vector allows tetracycline-regulated expression of the gene of interest from the strong CMV promoter in mammalian host cells expressing the Tet repressor (TetR) from the pcDNA 6/TR vector.

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

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