FuGENE HD Transfection Reagent
點擊次數(shù)：3021 更新時間：2010-11-24
FuGENE® HD Transfection Reagent is a next-generation transfection reagent, free of animal-derived components. It is designed to transfect a wide range of eukaryotic cells including insect cells and many cell lines not transfected well by other reagents （e.g ., MCF-7, RAW 264.7, PC-3, HeLa, MA-10, HepG2, SH-SY5Y, A7r5, STO, SCC-61, STSAR-90, SQ20B, T98）.
Note: For the most up-to-date list of cell types that have been successfully transfected with FuGENE® HD Transfection Reagent, visit our Transfection Special Interest Site at ａnd view our Transfection Reference Database. This site also contains much more information about the use of this reagent.
For life science research only.
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Generates meaningful physiological results, since FuGENE® HD Transfection Reagent has low cytotoxicity and doesn't "shock" cells （Figure 1）.
Figure 1: Comparison of transfection efficiency using two transfection reagents under standard medium conditions. After they were transfected with a plasmid carrying LacZ, cells were incubated for an additional 24 hours. β-Galactosidase expression was detected with theβ-gal StainingSetfrom Roche.
Figure 2: Comparison of transfection efficiency with two transfection reagents in the presence of higher serum concentrations. Standard growth medium was replaced by * FBS 1 hour prior to transfection. After cells were transfected with a plasmid carrying LacZ, they were incubated for an additional 72 hours. Expression of β-Galactosidase was detected with theβ-gal StainingSet from Roche.
- Provides a good model system for mimicking biological conditions, since FuGENE® HD Transfection Reagent can transfect cells in high （up to *） serum.
Figure 3: Transfection procedure with FuGENE® HD Transfection Reagent. Please note that the procedure does not require a media change after step 2.
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Generates high levels of protein expression in many adherent and suspension-adapted eukaryotic cell lines , including HEK-293 （Figure 4a）, Hep G2, CHO-S, CHO-K, COS-7, COS-1, and insect cell lines such as High Five （Figure 4b） and Sf9.
Figure 4a: High transfection efficiency and protein expression in suspension-adapted mammalian cells.
Panel a: HEK-293 EBNA suspension-adapted cells were transfected with plasmid DNA expressing GFP. The transfection reagent / DNA ratios used were 7:2, 6:2, 5:2, 4:2, and 3:2.
Panel b: The percentage of cells transfected was determined 28 hours post-transfection, and the quantity of GFP protein was estimated on a Coomassie-stained gel at 72 hours post-transfection.
* estimated yield
Figure 4b: High transfection efficiency and protein expression in High Five insect cells.
Panel a: Cells were transfected with a vector containing the LacZ gene. Transfection efficiency was demonstrated 24 hours post-transfection by histochemical staining for β-galactosidase activity （β-Gal Staining Set）
Panel b: Cells were transfected with vectors expressing human kinases Akt1 and MKK6. Protein expression in cell lysates was measured by western blot at 72 hours post-transfection.
- Saves time and minimizes mistakes, since transfection procedures with
- Simplifies downstream protein purification, since transfections with FuGENE® HD Transfection Reagent often produce high levels of expressed protein.
- Produces protein in insect cells much faster than the baculovirus cell expression system. Protein generating procedures that start with FuGENE® HD Transfection Reagent can take up to 18 fewer days （from transfection to protein purification） than the baclovirus system （Figure 5）.
Figure 5: Comparison of the traditional baculovirus cell expression method with an expression system that uses FuGENE® HD Transfection Reagent.
FuGENE® HD Transfection Reagent take very few steps （Figure 3） and do not need to be customized for different cell types.since FuGENE® HD Transfection Reagent works efficiently in both standard cell culture media （Figure 1） and those that contain up to * serum （Figure 2）, eliminating the need for media changes （either before or after transfection）. Composition: unique and proprietary blend of many components, in 80% ethanol.
Storage: stable at +2°C to +8°C.
Stability: 2 years, starting from date of manufacture.
Solution in 80% ethanol, filter-sterilized （0.1 µm filter）, packaged in transparent glass vials.
Lot-specific analysis:
Each lot of FuGENE®HD Transfection Reagent is carefully checked, using ISO DIN 9001-certified procedures, to assure product is performing according to specifications and is consistent from lot to lot. Functional analysis:
Cos 7 cells are transfected in 96 well plates with a reporter gene vector DNA, using 0.3 µl FuGENE® HD and 0.1 µg DNA or 0.25 µl FuGENE® HD and 0.1 µg DNA. Reporter gene activity is monitored via a colorimetric test against a reference lot. Activity must be ≥ 75% compared to a reference lot in at least one of the tested amounts, listed above.
Cytotoxicity analysis:
Cell growth is assessed using the Cell Proliferation Reagent WST-1assay which is an indirect measure of cytotoxicity. The remainder of cells from functional analysis is tested for viability.
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