TransMessenger Transfection Reagent is the first reagent specifically developed for transfection of cells with RNA. TransMessenger Reagent provides a fast and easy procedure and high transfection efficiencies (see figure "TransMessenger Reagent with HeLa Cells"). The reagent is a lipid-based formulation that is used in conjunction with a specific RNA-condensing enhancer and an optimized buffer. RNA molecules are condensed by the enhancer and then coated by TransMessenger Reagent for efficient transfer into eukaryotic cells. Strict quality control is performed to test for absence of RNase activity, lot-to-lot consistency, and low endotoxin levels (≤10 EU/ml). Our rigorous standards eliminate reagent variables that can adversely affect the efficiency of RNA transfection.
TransMessenger Reagent with HeLa Cells
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Expression of green fluorescence protein (GFP) in HeLa-S3 cells. 8 x 104 cells were seeded into 48-well plates and transfected 24 h later with 0.5 µg of an in vitro-transcribed GFP-encoding RNA (transcribed from P7ASP-GFP/Mlu) using 1 µl Enhancer R and 2.5 µl TransMessenger Reagent. Cells were analyzed 24 h post-transfection by fluorescence microscopy. Approximately 50% of the cells were successfully transfected. (P7ASP-GFP/Mlu kindly provided by J. Bogenberger, Stanford University Blood Center, Palo Alto, CA, USA.)
All TransMessenger Reagent components are provided as ready-to-use solutions. To generate TransMessenger–RNA transfection complexes, simply mix your RNA with Enhancer R and Buffer EC-R and incubate for 5 minutes at room temperature, then add TransMessenger Reagent and incubate for a further 5–10 minutes. The complexes are mixed with medium and added directly to the cells. Following a 3 hour incubation, the medium is changed and the cells are incubated until they are ready for analysis.
Optimal transfection results are achieved using high-purity RNA that is free of DNA, proteins, and other contaminants. RNA purified with RNeasy and Oligotex mRNA Kits is highly recommended. Since the amount of RNA is a critical factor for successful transfection, we recommend optimizing the amounts of RNA and TransMessenger Transfection Reagent for every cell type–RNA combination (see figure "Amount of RNA and TransMessenger Reagent vs. Transfection Efficiency"). To facilitate this, the reagent is provided with guidelines for optimization and starting points for optimization in different cell-culture formats.