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L3287
Escort™ IV Transfection Reagent
Lipid reagent for transient and stable transfection of mammalian and insect cells.
Synonym(s):
Gene delivery
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About This Item
UNSPSC Code:
41106502
NACRES:
NA.25
Pricing and availability is not currently available.
Recommended Products
grade
for molecular biology
Quality Level
form
liquid (aqueous solution)
usage
mL sufficient for 160-500 transfections
concentration
1 mg/mL
technique(s)
transfection: suitable
storage temp.
2-8°C
Related Categories
General description
Escort™ IV is a unique formulation of a proprietary polycationic lipid and a neutral non-transfecting lipid. This liposome-forming compound is used for transfection of nucleic acids into a wide variety of eukaryotic cell types.
Application
Suitable for transient and stable transfection of nucleic acids into cultured eukaryotic cells. Use approximately 4-16 μL Escort™ IV and 2 μg DNA per 6 cm cell culture plate. Protocol optimization provides very efficient transfection. For a list of cells that have been successfully transfected using Escort™ IV, see the Transfection Reagent Selection Guide.
Features and Benefits
- Suitable for stable and transient transfection
- Optimized for a wide variety of cell lines
- Low toxicity
- Compatible with both serum and serum-free transfection protocols
- Ideal for Sf9, Sf21 and S2 insect cells
Components
Escort™ IV formulation:
1 mg/mL total lipid in water
Note the identity of the lipids used in Escort™ IV is confidential.
1 mg/mL total lipid in water
Note the identity of the lipids used in Escort™ IV is confidential.
Caution
Do not freeze.
Principle
A stable complex is formed when Escort™ IV is mixed with DNA in the absence of serum. The complexes are stable and can be directly added to the cell culture medium, where they fuse with the cell membrane, releasing the DNA into the cytoplasm. Note: complex formation is inhibited by serum, but once stable complexes have formed, the presence of serum is without consequence.
Legal Information
Escort is a trademark of Sigma-Aldrich Co. LLC
related product
Product No.
Description
Pricing
Storage Class Code
10 - Combustible liquids
WGK
WGK 3
Flash Point(F)
Not applicable
Flash Point(C)
Not applicable
Personal Protective Equipment
dust mask type N95 (US), Eyeshields, Gloves
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What is the Department of Transportation shipping information for this product?
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Is optimizing the transfection protocol important?
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For many common cell lines, transfection reagent efficiency is very high and the protocols will not require any optimization. For hard-to-transfect cells or those ultimately expressing a toxic protein, the protocol should be optimized for best transfection efficiency. Taking time to optimize will give you more transfected cells with each procedure, which can mean more protein expressed and results that are reproducible.
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How can I increase the efficiency of my transfection?
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Transfection efficiency is affected by many different things, including plasmid size and purity, media components present, transfection reagent selected, amount of DNA and transfection reagent used, cell density, etc. Optimizing the protocol with respect to these concerns will allow you to achieve a higher transfection efficiency. For many cell lines and transfection reagents, optimized protocols are already available.
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How can I determine the efficiency of my transfection?
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Calculating transfection efficiency is very useful when optimizing transfection protocols. Transfection efficiency can be performed using a GFP-expressing plasmid. After transfection, cells are stained with propidium iodide and counted. The propidium iodide provides a count of the total cells in the population, and the GFP-expressing cells provide a count of the number of cells transfected. The transfection efficiency (%) can then be calculated by:(# GFP-expressing cells / total cell #) * 100
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What quality does the DNA need to be in order to use it for transfection?
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The DNA needs to be good quality or it may cause the cells to lyse and/or they won't transfect efficiently. Plasmid DNA prepared with a column-based DNA purification kit is suitable for transfections. Sigma's GenElute™ Minprep, Midiprep and Maxiprep kits work well for DNA plasmid purification. After preparing the DNA, confirm the OD A260:A280 ratio is greater than 1.6 for use in plasmid transfections.
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What is the difference between stable and transient transfection?
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When the DNA enters the nucleus of the cell, the plasmid is replicated by the cell machinery (transient transfection). During this time, RNA is transcribed and protein translated until the plasmid DNA is lost after a few cell divisions. This expression of the plasmid DNA, mRNA, and protein is transient (temporary).In some cases, the plasmid DNA is integrated into the host cell genome. This is usually accompanied by forced expression using a selection antibiotic and sometimes a cloning step (to be sure all cells have the same integration site). Once the DNA is stable, the cell line can be frozen and used to express protein for many years. Clones may even be screened for those expressing the highest amount of protein.
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What are the differences between the two Escort™ products?
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Each Escort™ transfection Reagent is a different lipid formulation. These different formulations are more readily taken up by different cells, presumably by endocytosis.Escort™ III is a unique formulation of a proprietary polycationic lipid and a neutral, non-transfecting lipid.Escort™ IV is a a confidential lipid.
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Is low cell passage number an important consideration for transfection?
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Yes, we recommend cells are at a low passage when being used for any application, including transfection. The reason why depends on what type of cells they are. Primary cells will undergo a finite number of divisions, and as they get closer to senesence they divide more slowly - both affecting their ability to take up DNA (transient transfection), and minimizing their abillity to incorporate the DNA into the genome (stable selection).Cultured common cell lines are often immortalized, and generally continue to aquire mutations, leading to a heterogenous population that may perform differently from cells of lower passage number - leading to results that are not reproducible.
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Can antibiotics be present in the medium during transfection?
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We recommend that no antibiotics are present during transfection. The process of transfection can make the cells somewhat more porous to allow for efficient DNA entry. During this time, antibiotics will also enter the cells more easily and the cells may show increased cell death. Wait until about 24 hours after transfection to resume the use of preventative antibiotics and/or start the use of selective antibiotics.
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What is transfection efficiency?
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Transfection efficiency is a measure of how many cells take up the DNA during the transfection process. Many transfection reagents can achieve a transfection efficiency of >90% in common cell lines. Other cell lines are hard to transfect, and require special reagents and/or techniques to achieve even a small population of transfected cells.
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