Description
TransMessenger Transfection Reagent is a ready-to-use, lipid-based reagent for RNA transfection of eukaryotic cells. Transfection of cells with RNA rather than DNA offers new possibilities for transfection experiments. Cell lines successfully transfected with TransMessenger Reagent include 293T, Jurkat, and Vero. This product can also be used for RNA transfection of primary human fibroblasts, rat cardiomyocytes, and rat neuronal cells.
TransMessenger Reagent provides a fast and easy procedure and high transfection efficiencies (see figure "TransMessenger Reagent with HeLa cells"). 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 together with starting points for optimization in different cell-culture formats. TransMessenger Transfection Reagent is highly suited for use in functional cardiac cell studies using neonatal rat ventricular cardiac cells. Cardiomyocytes from cryopreserved, dissociated neonatal rat ventricular cardiac cells (R-CM-561 [QBMCellScience.com]) display excellent viability, pharmacology, morphology, and connectivity, as well as contractile and electrical activity necessary for use in functional screening (see video Transfection control: Functional Cardiomyocytes Beating). To evaluate the suitability of using TransMessenger Transfection Reagent in functional cardiac cell studies, cells were transfected with a GFP-expression vector 4 hours after plating, and were fixed 5 days post-transfection. Use of TransMessenger Transfection Reagent resulted in an excellent, 60% survival rate post-transfection with a transfection efficiency of 10–13%. All cardiomyocytes displayed beating post-transfection (sees videos GPF-Positive Cardiomyocytes Beating Post-Transfection and Functional Cardiomyocytes Beating Post-Transfection), indicating functional cells. The number of transfected cardiomyocyte versus non-cardiomyocyte cells was evaluated and results demonstrate that a high number of cardiomyocyte and non-cardiomyocyte cells were transfected. Equivalent morphologies between transfected and non-transfected cells were also observed (see figure Cardiac cell studies).
TransMessenger Transfection Reagent is the first reagent specifically developed for transfection of cells with RNA. 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.
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 hours 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 is highly recommended.
