What are 293f cells?

What are 293f cells?

293-F cells are GIBCO brand cells. These are clonal isolates derived from transformed human embryonal kidney (HEK) cells. The 293-F strain is a fast-growing variant of the 293 cell line. They have been adapted to suspension culture in serum-free media, 293 SFM II.

What are FreeStyle 293-f cells?

FreeStyle 293-F cells are derived from HEK293 cells; they were just cloned extensively for growth, viability, and protein production in the new serum-free FreeStyle 293 medium.

What is the difference between 293T and 293 ft?

You can use HEK293 cells for lentivirus production, but keep in mind that the titers will be lower. 293T cells and 293FT cells grow faster and are easier to transfect than HEK293 cells and result in higher titers of lentivirus. So most lentiviral protocols use 293FT cells to produce lentivirus.

What are CHO cells and what are they used for?

CHO cells are the most common mammalian cell line used for mass production of therapeutic proteins. They can produce recombinant protein on the scale of 3-10 grams per liter of culture.

Does lentivirus lyse cells?

The cells should not lyse with lentivirus. The virus is secreted into the media. An important consideration is that the expression levels in your target cells is not high enough to get a clear signal.

How do 293T cells make virus?

During incubation, the 293 cells transcribe the DNA of interest from the lentiviral plasmid, creating an RNA copy of the lentiviral construct. They also transcribe and translate the genes in the packaging vectors into viral proteins.

Are CHO cells cancerous?

CHO cells can also be kept in suspension cultures; in contrast to cancer cells, they are genetically stable; they can be reproduced with expression vectors that contain the “gene of interest” (GOI); they can be transfected; and they remain stable during the process of selection, amplification, single-cell cloning and …

Why do we use CHO cells?

CHO cells are the most common mammalian cell line used for mass production of therapeutic proteins. Products of CHO cells are suitable for human applications, as they allow post-translational modifications to recombinant proteins which can function in humans.

How long does it take for lentivirus to infect cells?

All lentiviral vectors present in the transduction mix need at least 5 hours to penetrate the cells of interest. Based on the experiment, the transduction can be left from 5 hours to an overnight incubation.

Can lentivirus infect mouse cells?

Yes, you can use lentivirus to stably transduce rodent primary cells. Although lentiviral vectors do transduce non-dividing cells, the efficiency is not always as good as with dividing cells. High multiplicity of infections (MOI) may lead to multiple integrations and very high levels of gene expression.

How do you transduce a virus to a cell?

c. Transduction

1. Thaw the lentivirus on ice. Mix 8 µl Polybrene (1 mg/ml aliquot) with 957 µl culture.
2. The next day, exchange Lentivirus/Polybrene mixture by fresh culture medium. Incubate cells at standard cell culture conditions.
3. concentrations range from 0.1-10 μg/ml. Replace the culture medium 48-72 hours.

Can you use expifectamine293 on freestyle 293 cells?

You should be able to use the ExpiFectamine293 Reagent for transfection of Freestyle 293-F cells grown in FreeStyle 293 Expression Medium; however, the enhancers will provide only a little boost in expression as they are designed to work with higher density cultures that FreeStyle 293 Expression Medium cannot support.

Which is the best transfection reagent for 293 cells?

293fectin™ has been optimized for transfection of suspension 293 cells (e.g. FreeStyle™ 293-F cells, Cat. no. R790-07) in defined, serum-free FreeStyle™ 293 Expression Medium (Cat. no. 12338-018). Using 293fectin™ provides the following advantages:

How are Invitrogen freestyle 293 cells adapted to suspension culture?

The system includes Invitrogen FreeStyle 293-F cells that have been adapted to serum-free, suspension culture in Invitrogen FreeStyle 293 Expression Medium. Transfection and expression experiments may be performed directly in FreeStyle 293 Expression Medium without the need to change media.

How to determine the viability of 293fectin transfection?

On the day of transfection, determine the viability and the amount of cell clumping from a small aliquot of cells using the trypan blue dye exclusion method. Vigorously vortex for 45 seconds to break up cell clumps and determine total cell counts using a Coulter Counter or a hemacytometer. Viability of cells must be over 90%.