HepG2 Transfection

LIVER HEPATOCELLULAR CARCINOMA

HepG2 Cell Transfection

HepG2 Cell Transfection Kit

A pre-optimized HepG2 Transfection Kit is available from Altogen Biosystems, which includes:

  • HepG2 Transfection Reagent (0.5 ml / 1.5 ml / 8.0 ml)
  • Transfection Enhancer (0.5 ml), and
  • Complex Condenser (0.5 ml)

The transfection kit is optimized in HepG2 cells and enables a high efficiency transfer of siRNA, miRNA or plasmid DNA into the cells.  The transfection reagent enables either a forward or reverse transfection method in order to achieve at least 90% transfection efficiency.

Transfection Protocol

An optimized protocol to transfect HepG2 cells in a 24-well plate is described below:

  1. Plate 7,500-12,000 HepG2 cells per well in 0.5 ml of complete growth medium 12-24 hours prior to transfection
  2. Wash with 1xPBS and add 0.5 ml of fresh growth medium
  3. Prepare transfection complexes by mixing 40 µl of serum-free medium, 5.5 µl of transfection reagent, and (referred to a final volume including growth medium)
    • 750 ng DNA (or mRNA), or
    • 30 nM -50 nM of siRNA (or microRNA)
  4. Incubate transfection complexes at RT for 15-30 minutes
  5. Optional: Add 2 µl of Complex Condenser. This reagent increases transfection efficiency by reducing the size of transfection complex; however, it may increase cell toxicity
  6. Add prepared transfection complexes to 0.5 ml of complete growth medium with HepG2 cells (from step 2)
  7. Incubate cells at 37ºC in a humidified CO2 incubator
  8. Assay for phenotype or target gene expression 48-72 hours after transfection

Optional: Adding Transfection Enhancer reagent can increase transfection efficiency. Add 2 µl of Transfection Enhancer reagent 12-24 hours after transfection

If the viability of HepG2 cells being transfection is affected 16-24 hours post-transfection, changing the growth medium and eliminating redundant exposure of cells to transfectant can reduce the level of cytotoxicity

HepG2 Research Studies

HepG2 cells are useful as a model system in vitro for human hepatic cancer cells. When cultured properly, they differentiate, forming apical and basolateral cell surfaces, resembling in vivo liver structures. This differentiation makes them very helpful in the study of intracellular transport of molecules between the different liver cell types, dysfunction of which can cause several human liver diseases including progressive familial intrahepatic choestasis and Dubin-Johnson Syndrome. This cell line is also useful to study toxicity of substances to the liver, as well as for the examination of liver metabolism. HepG2 cells are also important in the study of drug metabolism, as many toxic metabolites are formed in the liver and being able to identify these in vitro reduces the need for in vivo studies. In addition, HepG2 cells have been used in trials with artificial liver devices.

  • A study of gene expression between HepG2 carcinoma cells and normal hepatocyte cells: This study compared the regulation of genes in two types of cells to determine if there was a “gene signature” for hepatoma cells. This identification allows future research in the field to focus on genes which have been identified as related to the carcinoma genotype. [LINK]
  • A comparison of the assembly and secretion of very low density lipoproteins (VLDL) between HepG2 cells and Huh-7 cells: VLDL secretion is often studied in HepG2, but HepG2-produced VLDL is dense and lipid-poor compared with in vivo VLDL. The study found that HuH-7 cells did not offer any advantage over HepG2 cells in terms of apoB100 lipoprotein particle production. [LINK]
  • Magnetic nanoparticles targeting HepG2 cells: A team in Taiwan has engineered magnetic nanoparticles with functionalized surface ligands designed to enhance uptake of the particles by HepG2 cells. Confocal microscopy was used to visualize the particles and showed they were released from endosomes after entering the cells. These nanoparticles have been shown to be non-cytotoxic and may have in vivo applications. [LINK]
  • Folate deficiency and apoptosis in HepG2 cells: A study was completed to investigate the link between depriving liver cells of folate and the death rate of cells. It was determined that folate-deficient medium caused decreased growth and viability as well as an increased propensity towards apoptosis, independent of p53-mediated pathways. [LINK]

Vendors

HepG2 cell line is commercially available from American Type Culture Collection (ATCC) and DSMZ (German Resource Centre for Biological Material).

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