For 2500–6500 transfections in 60 mm dishes or 5000–10,000 transfections in 6-well plates.
The PolyFect Transfection Reagent is intended for molecular biology applications. This product is not intended for the diagnosis, prevention, or treatment of a disease.
High transfection efficiencies in CHO and 293 cells.
Transfection efficiencies for PolyFect Reagent and 2 lipid-based reagents were compared. Cells cultured in 6-well plates were transfected with a beta-galactosidase-reporter plasmid. The appropriate protocol was used for transfection with PolyFect Reagent. Optimized protocols were used for transfection with Reagents L2 and F; these were: 10 µl Reagent L2 with 3 µg DNA and 15 µl Reagent F with 2 µg DNA for CHO cells, and 10 µl Reagent L2 with 2 µg DNA and 7 µl Reagent F with 3 µg DNA for 293 cells. All transfections were performed in triplicate. Two independent transfection experiments are shown for 293 cells.
PolyFect transfection procedure.
The transfection procedure is fast and simple - just add PolyFect Reagent to the DNA solution, mix, incubate for 5-10 minutes, add growth medium (which can contain serum and antibiotics), and pipet the PolyFect-DNA complexes onto the cells. The cells are then incubated for gene expression.
Note the highly branched structure of an activated dendrimer in this schematic representation.
A model of the PolyFect-DNA complex shows PolyFect Reagent (purple balls) interacting with DNA (black) to form a ring-like (toroid-like) structure. The upper right section of the illustration shows naked DNA, the lower section shows the interaction between dendrimers and DNA inside the complex, and the upper left section shows the final complete coverage of DNA within the complex.
PolyFect Reagent with HeLa cells.
Expression of [A] beta-galactosidase and [B] green fluorescent protein (GFP) in HeLa cells is shown. Cells were cotransfected in 6-well plates with beta-galactosidase- and GFP-reporter plasmids using PolyFect Transfection Reagent and the HeLa cell protocol. Expression was visualized by X-gal staining or fluorescence microscopy 2 days post-transfection.
PolyFect Transfection Reagent is a solution of specifically designed activated-dendrimers. The reagent consists of dendrimer molecules of a defined spherical architecture with branches radiating from a central core (see figure "Activated dendrimer"). The branches terminate at charged amino groups, which can interact with negatively charged phosphate groups of nucleic acids. PolyFect Reagent assembles DNA into compact structures (see figure "PolyFect–DNA interaction"), that bind to the cell surface and are taken into the cell by nonspecific endocytosis. The reagent buffers the pH of the endosome, leading to pH inhibitition of endosomal nucleases, which ensures stability of PolyFect–DNA complexes.
PolyFect Transfection Reagent is provided as a ready-to-use solution. The transfection procedure (see flowchart "PolyFect transfection procedure") is fast and simple — just add PolyFect Reagent to the DNA solution, mix, incubate for 5–10 minutes, add growth medium (which can contain serum and antibiotics), and pipet the PolyFect–DNA complexes onto the cells. The cells are then incubated for gene expression. No post-transfection removal of complexes or medium change/addition is necessary, making the procedure fast and easy.
Transfection using PolyFect Reagent requires minimal handling and removal of transfection complexes is not required, making this reagent highly suitable for high-throughput screening. PolyFect Reagent provides outstanding transfection efficiencies, excellent reproducibility, and minimal cytotoxicity in high-throughput transfection, and is available in bulk quantities. For optimized protocols for transfection of COS-7, NIH/3T3, HeLa, HeLa-S3, 293, and CHO cells in 96-well plates, please contact QIAGEN Technical Services.
PolyFect Transfection Reagent is highly suited for efficient transfection of standard cell lines in:
Studies on gene expression and function
Drug discovery and development studies
Plasmid transfection, protein overexpression
COS-7, NIH/3T3, HeLa, 293, CHO cells
Optimized protocols for COS-7, NIH/3T3, HeLa 293, and CHO cells. Transfection in the presence of serum
Number of possible transfections
Up to 50–100 transfections in 6-well plates / 1 ml reagent