High-Density Mammalian Transient Protein Expression
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The Expi293™ System gives you the benefit of a stable cell system in much less time. It’s easy to produce 3–5 times more protein with much less effort.
The Expi293™ Expression System is a major advancement in transient expression technology for rapid and ultra high-yield protein production in mammalian cells. It is based on high density culture of Expi293F™ cells in Expi293™ Expression Medium. Transient expression is powered by the new cationic lipid-based ExpiFectamine™ 293 transfection reagent in combination with optimized transfection enhancers designed to work specifically with this transfection reagent. All components work in concert to generate 2- to 10-fold higher protein yields than conventional culture systems such as the FreeStyle™ 293 Expression System. Expression levels of greater than 1 g/L were achieved for IgG and non-IgG proteins (See Figure 1).
Figure 1. Expression of human IgG and Fc-tagged Cripto achieve expression levels of over 1g/L in the Expi293™ Expression System.
What’s in the System?
The components included in the kit are:
- 2 vials of frozen Expi293F™ cells
- 1 liter of Expi293™ Expression Medium
- 1 ExpiFectamine™ 293 Transfection Kit, sufficient to transfect 1 liter of culture
- OptiMEM® Reduced Serum Medium
- An Antibody-Expressing Positive control
The ExpiFectamine™ 293 Reagent and ExpiFectamine™ 293 Transfection Enhancers 1 and 2 are sold together as the ExpiFectamine™ 293 Transfection kit. All other components of the system are available separately.
What Makes Expi293™ Expression System Unique?
Expi293™ Expression Medium is a chemically defined, serum-free, protein-free medium. Expi293™ Expression Medium enables high density cell culture (See Figure 2) and transfection, allowing for significantly higher volumetric protein yields than traditional culture systems.
Figure 2. Cell viability and viable cell density over time for cells grown in Expi293™ Expression Medium vs. other 293 culture media.
Expi293F™ cells are specifically adapted to exhibit higher pg/cell/day productivity than typical 293 cells and produce up to 1.7x more protein than Freestyle™ 293F cell when used in the Expi293™ Expression System.
ExpiFectamine™ 293 transfection reagent is capable of high-efficiency transfection of high-density 293 cultures, and is coupled with transfection enhancers that further boost transfection performance and expression levels. (See Figure 3).
Figure 3. Expression of hIgG, with and without transfection enhancers.
How Does the Kit Work?
The Expi293™ Expression System contains all the components needed to easily produce high levels of recombinant protein in your own laboratory. The only additional component required is a mammalian expression vector containing the DNA for your protein of interest.
Expi293F™ cells are thawed and grown in suspension culture in Expi293™ Expression Medium. Cells are typically grown in flasks on a shaker platform inside a CO2 cell culture incubator or in bioreactors. The cells are transfected using ExpiFectamine™ 293 Transfection Reagent. Transfection enhancers 1 and 2 are then added to the culture 16-18 hours post-transfection. There is no need to change culture medium or otherwise supplement the cultures during protein production. Protein harvest can be performed from 2 to 7 days post-transfection, depending on the expression profiles of the protein of interest. For expression of recombinant antibodies, maximum yields are often seen from 5 to 7 days post-transfection.
The system is highly scalable and should produce similar volumetric protein yields in transfection formats ranging from 1 mL cultures in a 24-well plate up to 1L cultures in shake flasks (See Figure 4).
Figure 4. Expression of Fc-tagged Cripto achieves similar volumetric protein yields at 3 different scales.
The Expi293™ Expression System is a rapid method of producing mg to gram quantities of recombinant protein in mammalian cells. For the expression of human or other mammalian proteins, expression in the human-derived 293 cells offers the benefit of more native folding and post-translational modifications, such as glycosylation, when compared to expression systems based on hosts such as E. Coli, yeast, or insect cells.
*Protein yields in each culture are expressed as normalized values, based on the amount obtained using FreeStyle™ Expression Medium supplemented with PEI in 1 L of culture.
- Adaptation of Expi293™ to the WAVE Bioreactor™ System
- Expi293™ MembranePro™ Expression System App Note
- The Expi293™ Expression System—Optimizing protein yield
- Expi293™ Expression System—Optimizing culture conditions to maximize protein yield and enable use of 96-well microtiter plates
- Using the Expi293™ Expression System under animal origin–free (AOF) conditions