Bio-Rad TransFectin™ Lipid Reagent - Manual

TransFectin ™ Lipid Reagent
0.5 ml 170-3350
1.0 ml 170-3351
5 x 1.0 ml 170-3352
For Research Use Only
Store at 4°C
Storage and Stability
TransFectin Lipid Reagent is shipped on ice. Store at 4º C upon receipt. Do not store below 0ºC. TransFectin is
stable for 6 months from date of purchase when stored at 4º C.
Contents
TransFectin contains either 0.5 ml (Catalog Number 170-3350), 1.0 ml (Catalog Number 170-3551), or 5 x 1.0 ml
(Catalog Number 170-3352) of a proprietary lipid formulation. One milliliter is generally sufficient for 125 to 200 cell
transfections in 35 mm plates. TransFectin comes as a 1.5 mg/ml solution.
Reagent
TransFectin Lipid Reagent is a mixture of a proprietary cationic compound and a co-lipid (1,2-Dioleoyl-sn-Glycero-3-
Phosphoethanolamine). These compounds have been optimized for intracellular delivery of nucleic acids into cultured
mammalian cells in the presence of serum at cell densities from 50% to greater than 90%. For most cell
lines, high levels of expression can be obtained using concentrations of TransFectin and DNA suggested in this manual
for given sizes of plates/wells. If presently using a cationic lipid for transfection, start using TransFectin at current
conditions and also try reducing and increasing the volume of TransFectin from 25 to 50% of current amount. For best
results it is important to determine the optimal amount of DNA and lipid for any given cell line.
Recommendations for Best Results
• For most cell lines use a ratio of DNA (µg) to lipid (µl) of 1:2–1:3 as a starting point to optimize conditions.
• Invert the tube to mix contents before using.
• TransFectin is designed to work in media containing serum but can be used in the absence of serum.
• Use sterile polystyrene plastic ware (e.g. 12 x 75 mm tubes or multi-well trays) to prepare the plasmid solutions
and lipid solutions. Polystyrene is recommended because cationic lipid-plasmid complexes may bind to
polypropylene.
Optimization
Determining the optimum conditions for transfection efficiency is essential for maximizing gene expression
and minimizing cellular toxicity. The two most important parameters to optimize for any given culture vessel
and cell density are the concentration of TransFectin and the amount of nucleic acid. See Table 1 for
suggested reagent and DNA concentrations for different culture vessel sizes. In general,
gene expression will increase, plateau and then decrease with increasing concentrations of TransFectin.
This decrease in gene expression correlates with reduced cell viability. As increasing amounts of plasmid
are added to the cells, gene expression will increase, then plateau or even decrease since the amount of
DNA used can directly affect toxicity. If toxicity is encountered, try reducing lipid or DNA amounts used in
transfection. The concentration of TransFectin and the amount of plasmid required for maximal expression
may vary from one cell line to another.
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