GFP expression in A-375 (Melanoma) cells, 48 hours after transfection by FuGENE®HD and Competitor Lipofectamine® 2000, 10,000 cells/well.

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 GFP Expression  Cell Morphology

Reagent / DNA Ratio
0.3μl/0.1μg
8μl Complex per well
Complex Incubation 10 min

FuGENE®HD

 


Reagent / DNA Ratio
0.3μl/0.1μg
8μl Complex per well
Complex Incubation 10 min

Reagent / DNA Ratio
0.5μl/0.2μg
50μl Complex per well
Complex Incubation 20 min

Lipofectamine® 2000

 


Reagent / DNA Ratio
0.5μl/0.2μg
50μl Complex per well
Complex Incubation 20 min

Reagent / DNA Ratio
0.3μl/0.1μg
8μl Complex per well
Complex Incubation 10 min

FuGENE®HD

 


Reagent / DNA Ratio
0.3μl/0.1μg
8μl Complex per well
Complex Incubation 10 min

Reagent / DNA Ratio
0.5μl/0.2μg
50μl Complex per well
Complex Incubation 20 min

Lipofectamine® 2000

 


Reagent / DNA Ratio
0.5μl/0.2μg
50μl Complex per well
Complex Incubation 20 min
 Transfection with competitor reagent was performed according to manufacturer’s instructions

Protocol for A-375 Cell Transfection by FuGENE HD with gWIZ™ GFP plasmid.

  1. Cell plating

    A-375 cells were seeded from 90-100% confluent culture the day before transfection with the density 15,000 cells per well in 100 μl complete growth medium DMEM with 1.5 g/L sodium bicarbonate +10%Fetal Bovine Serum in 96-well plates. A-375 cells were at approximately 80% confluence on a day of transfection.

  2. Complex preparation (per 20 wells)

    Tissue culture 96-round bottom well plates were used for complex preparation:

    1. Prepare 0.0125μg/μl gWIZ™ GFP DNA solution in OptiMEM®.
    2. Add 6 μl of reagent to 160 μl of DNA solution.
    3. Mix carefully by pipetting (15 times).
    4. Incubate 10 min at room temperature.
    5. Add 8 μl of complex per well to the cells, and mix thoroughly.
  3. Incubation

    Place the cells into CO2 incubator for 48 hours.

  4. Detection of GFP expression

    Monitor the transgene expression with fluorescent microscope or flowcytometer.