Protocol- 50% efficiency in hepatocyte-like cells and primary hepatocytes
- Gentle to cells
- Easy-to-use protocol
- Compatible with serum and antibiotics
jetPEI™-Hepatocyte is a DNA transfection reagent designed to transfect Hepatocyte-like cells. jetPEI™-Hepatocyte is recommended to transfect primary hepatocytes and cell lines such as human hepatocarcinoma HepG2 and murine hepatocytes BNL-CL.2.
INTELLECTUAL PROPERTY:
The use of polyethylenimine (PEI) or polypropylenimine (PPI) or cationic polymers similar in structure thereto for transfecting cells, as well as compositions comprising these cationic polymers and at least one nucleic acid, are the subject matter of U.S. Patent No. 6,013,240, EP Patent No. 0770140 and foreign equivalents, for which Polyplus-transfection™ is the worldwide exclusive licensee.
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Complexes formation is perfectly performed with NaCl solution. 0.5 ml of jetPEI™-Hepatocyte is sufficient to perform 160 transfections in 24-well plates. |
Compared to our standard jetPEI™ transfection reagent, jetPEI™-Hepatocyte provides significantly higher transfection efficiencies on human hepatocarcinoma cells HepG2 as well as on the difficult-to-transfect primary human hepatocytes (Fig. 1).
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Figure 1. Comparative efficiency of jetPEI™-Hepatocyte versus jetPEI™. HepG2 (50,000 cells in 24-w) and primary human hepatocytes (100,000 cells in 24-w) were transfected using 3.2 µl of both reagents with 1 µg of pCMVEGFPLuc, in 1 ml of complete medium containing 10% of serum. The luciferase activity was determined 48h post-transfection. Transfection efficiency was expressed as relative efficiency obtained with jetPEI™ compared to jetPEI™-Hepatocyte. |
jetPEI™-Hepatocyte protocol is as simple as that of the jetPEI™: Mix DNA with the reagent to form complexes and simply add the mixture to cells. jetPEI™-Hepatocyte is compatible with serum1 and antibiotics. No media changes or washes are required. Transgenic protein expression is determined 24 h to 72 h post-transfection.
Primary human hepatocytes 24 h and 48 h after transfection using jetPEI™-Hepatocyte of the luciferase plasmid pCMVEGFPLuc show 50% transfection efficiency. In addition, cells look healthy (Fig. 2).
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Figure 2. Primary human hepatocytes expressing GFP 24 h and 48 h after transfection using jetPEI™-Hepatocyte. Primary human hepatocytes (100,000 cells in 24-w) were transfected with 1 µg of pCMVEGFPLuc at N/P ratio of 8, in 1 ml of complete medium containing 10% of serum. Cells were visualized by fluorescence microscopy. |
The most popular hepatocyte-derived cell lines BNL-CL.2 and HepG2 were successfully transfected using jetPEI™-Hepatocyte and commonly show approximately 50 % and 30% transfection efficiency, respectively (Fig. 3 and 4).
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Figure 3. BNL-CL.2 cells expressing ßgalactosidase after transfection using jetPEI™-Hepatocyte. BNL-CL.2 cells (50,000 cells in 24-w) were transfected using 3.2 µl of reagent with 1 µg of pCMVEGFPLuc, in 1 ml of complete medium containing 10% of serum. ?-galactosidase gene expression was visualized by X-gal staining after 24h. |
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Figure 4. HepG2 cells expressing GFP 72 h after transfection using jetPEI™-HepatocyteHepG2 cells (50,000 cells in 24-w) were transfected using 3.2 µl of reagent with 1 µg of pCMVEGFPLuc, in 1 ml of complete medium containing 10% of serum. Cells expressing GFP protein were visualized by fluorescence microscopy. |
jetPEI™-Hepatocyte is a galactose-bearing linear polyethylenimine designed to enhancethe transfection of cells expressing galactose-specific membrane lectins, such as hepatocytes that express the asialoglycoprotein receptor (ASGP-R or Gal/GalNAc receptor). Cell targeting is the result of binding of the galactose residues to the specific cell-surface receptors, leading to internalization of the DNA complexes. Galactose-bearing PEI has also been used to target oligonucleotides to hepatocytes in cell culture2.
1. Zanta M. A., O. Boussif, A. Adib and J. P. Behr (1997) In-Vitro Gene Delivery to Hepatocytes with Galactosylated Polyethylenimine. Bioconj. Chem. 8, 839-844
2. Kren B. T., P. Bandyopadhyay and C. J. Steer (1998) In-Vivo Site-Directed Mutagenesis of the Factor-IX Gene by Chimeric RNA/DNA Oligonucleotides. Nature Med 4, 285-290
