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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L3287
Escort™ IV Transfektionsreagenz
Lipid reagent for transient and stable transfection of mammalian and insect cells.
Synonym(e):
Gene delivery
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| Größe/SKU | Verfügbarkeit | Preis |
|---|---|---|
1 mL | Warenkorb auf Verfügbarkeit prüfen | € 327,00 |
Über diesen Artikel
NACRES:
NA.25
UNSPSC Code:
41106502
Form:
liquid (aqueous solution)
Grade:
Molecular Biology
Technique(s):
transfection: suitable
Concentration:
1 mg/mL
€ 327,00
Warenkorb auf Verfügbarkeit prüfen
Technischer Dienst
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Unterstützung erhaltengrade
Molecular Biology
Quality Segment
form
liquid (aqueous solution)
usage
mL sufficient for 160-500 transfections
concentration
1 mg/mL
technique(s)
transfection: suitable
storage temp.
2-8°C
General description
Escort™ IV ist eine spezielle Formulierung aus einem proprietären polykationischen Lipid und einem neutralen, nicht transfizierenden Lipid. Diese liposomenbildende Verbindung wird für die Transfektion von Nukleinsäuren in eine Vielzahl von eukaryotischen Zelltypen verwendet.
Application
Geeignet für die transiente und stabile Transfektion von Nukleinsäuren in kultivierte eukaryotische Zellen. Ungefähr 4–16 μL Escort™ IV und 2 μg DNA pro 6-cm-Zellkulturplatte verwenden. Die Optimierung des Protokolls sorgt für eine sehr effiziente Transfektion. Eine Liste der Zellen, die mit Escort™ IV erfolgreich transfiziert wurden, finden Sie im Transfection Reagent Selection Guide.
Biochem/physiol Actions
Wenn Escort™ IV mit DNA in Abwesenheit von Serum gemischt wird, bildet sich ein stabiler Komplex. Die Komplexe sind stabil und können direkt in das Zellkulturmedium gegeben werden, wo sie mit der Zellmembran verschmelzen und die DNA in das Zytoplasma freisetzen. Hinweis: Die Komplexbildung wird durch Serum gehemmt, aber sobald sich stabile Komplexe gebildet haben, spielt die Anwesenheit von Serum keine Rolle.
Features and Benefits
- Für stabile und transiente Transfektionen geeignet.
- Für eine breite Palette von Zellarten optimiert.
- Geringe Toxizität
- Kompatibel mit serumhaltigen und serumfreien Transfektionsprotokollen
- Ideal für Sf9-, Sf21- und S2-Insektenzellen
Other Notes
Formulierung von Escort™ IV:
1 mg/mL Gesamtlipid in Wasser
Hinweis: Die Identität der in Escort™ IV verwendeten Lipide ist vertraulich.
1 mg/mL Gesamtlipid in Wasser
Hinweis: Die Identität der in Escort™ IV verwendeten Lipide ist vertraulich.
Legal Information
Escort is a trademark of Sigma-Aldrich Co. LLC
Disclaimer
Nicht einfrieren.
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Dieser Artikel | |||
|---|---|---|---|
| description Lipid reagent for transient and stable transfection of mammalian and insect cells. | description Lipid reagent for transfecting sensitive and primary cells | description Animal-free polycationic liposomal transfection reagent optimized for the transfection of HEK293 cells grown in suspension culture. | description >99% (TLC), liquid, suitable for transfection |
| grade Molecular Biology | grade Molecular Biology | grade - | grade - |
| technique(s) transfection: suitable | technique(s) transfection: suitable | technique(s) transfection: suitable | technique(s) transfection: suitable |
| form liquid (aqueous solution) | form liquid (aqueous solution) | form liquid | form liquid |
| Quality Level 100 | Quality Level 200 | Quality Level 200 | Quality Level - |
| concentration 1 mg/mL | concentration 1 mg/mL | concentration - | concentration - |
| storage temp. 2-8°C | storage temp. 2-8°C | storage temp. 2-8°C | storage temp. 2-8°C |
Lagerklasse
10 - Combustible liquids
wgk
WGK 3
flash_point_f
Not applicable
flash_point_c
Not applicable
ppe
Eyeshields, Gloves
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How can I increase the efficiency of my transfection?
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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 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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Why do I see a precipitate in my cell culture after lipid-based transfection?
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The precipitate is likely excess lipid or EDTA and will probablly not affect transfection efficiency. If your DNA plasmid is suspended in TE, be sure the concentration of EDTA is <0.3 mM, or suspend the DNA in sterile molecular biology grade water instead.
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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 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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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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Which Escort™ transfection reagent should I use for my application?
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Escort™ III is best for sensitive cells and primary cells, particularly PC-12 and Jurkat.Escort™ IV is excellent for mammalian cell lines, primary cells and insect cells.
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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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