L growth medium for a single well of a 24well plate. Lipofectamine 2000 transfection, the expression of gfp was detected in 76,4% of hela cells with a mean fluorescence intensity of 437. Transitx2 dynamic delivery system and lipofectamine 2000 transfection reagent were used to. Hepesbuffered physiological saline solution containing.
The optimized protocol for a 24well plate to transfect a549 cells is described below. Invitrogen lipofectamine 3000 transfection reagent 1. This reagent delivers superior transfection efficiency. Xfect provides excellent transfection for most cell types. One can get over 4050% transfection efficiency with plasmid dna and over 85% with sirna. Igur high efficiency transfection on a broad range. Each reagent was used to transfect hek 293, hela, lncap, a549, and. Introducing the versatile reagent with the power to. Singlecell transfection of adherent cells has been accomplished using singlecell electroporation scep with a pulled capillary. Transfection reagents are highly efficient for dna and sirna transfection. A preoptimized a549 transfection kit is available from altogen biosystems, which includes. A549, carcinomic human alveolar basal epithelial cells. I use 24well plates for transfections in triplicates. Hela cells typically transfect with 8590% efficiency.
Lipofectamine 2000 transfection reagent is a proprietary formulation for the transfection of nucleic acids dna and rna into eukaryotic cells and provides the following advantages. Transfection guide overview of transfection methods promega. On the day of transfection just prior to preparing complexes, plate 1. However, significant toxicity from lipofectamine 2000 was also reported in. This reference provides a recommended procedure to transfect plasmid dna into human a549 lung carcinoma cells atcc, cat. Increase transfection efficiency and cell viability with. How to maximize transfection efficiency in a549 cells. Enhanced transfection efficiency using lipofectamine 3000 reagent.
Transfection guide overview of transfection methods. A b on the day of transfection, which should be 1 day following cell plating, perform the following steps, which have been optimized for a single well of a 24well plate using invitrogen lipofectamine 3000 transfection. Lipofectamine 2000 transfection reagent dartmouth college. One day prior to transfection, adherent cells were plated onto 24well plates at 0. Transfecting plasmid dna into a549 cells using lipofectamine. Lipofectamine 3000 reagent maintains a high transfection efficiency within a robust dynamic range of lipid doses for quick and easy optimization. With proprietary shaved cell transfection protocol, polyjet left panel gives up to 70% gfp positive cells vs. Invitrogen lipofectamine 3000 transfection reagent. Figure 3 illustrates differences in expression and toxicity profiles for two common cell types, a549 and mdck cells. Transfection efficiencies lower than 50% were not reported in this table. Mirus lipofectamine 2000 and oligofectamine invitrogen. Transfection efficiency was assessed by facs analysis in various cell lines 24 h after transfection in 24well plates.
Copy link enlarge image comparison of forward and reverse transfection in a549. Endofectin transfection reagent, cell lines genecopoeia. Lipofectamine 2000 right panel around 5% efficiency. Cloned genes can be transfected into cells for biochemical characterization. Viafect reagent offers performance similar to or better than that of lipofectamine 2000 reagent, and it may be suitable. A new transfection reagent formulated specifically for a549 cells. Transfection protocol of adherent a549 cells 96well plate 1. On the day of transfection, which should be 1 day following cell plating, perform the following steps, which have been optimized for a single well of a 24well plate using lipofectamine 3000 transfection reagent. A549 dharmafect 2 hct116 dharmafect 1 huh7 dharmafect 3 a673 lipofectamine 2000. From expert transfection reagent developer molecular transfer inc. A549 transfection kit lung carcinoma, ccl185 altogen. A549 lipofectamine 3000 lipofectamine 2000 fugene hd lipofectamine 2000 lipofectamine 3000 competitor 1 competitor 2 western blot, hepg2 gststat. Different cell lines were transfected with a gfp reporter vector using the indicated transfection.
Viafect transfection reagent allows highefficiency transfection of a wide range of. Protocol for lipidmediated transient transfection in a549 epithelial lung cell line. I use lipofectamine 2000 invitrogen for transfection of both sirna and plasmid dna into these 293 cell lines. Transient plasmid dna transfection into hela and hacat. Lipofectamine 3000 and rnaimax outperformed lipofectamine 2000. Dna transfection for luciferase assay in human alevolar. Choosing the right transfection reagent for optimal efficiency. Lipofectamine 2000 cd reagent is a proprietary animal originfree formulation for transfecting nucleic acids into eukaryotic cells. In biontex experiments gfp positivity increased to 90. Transfection the delivery of dna or rna into eukaryotic cells is a powerful tool used to study and control gene expression. Altogen biosystems provides in vivo transfection reagents, over 100 preoptimized in vitro transfection kits for cell lines and primary cells, and electroporation delivery products. I have used 3 kinds of transfection reagents lipofectamine, geneporter2, jetpei to test which one was better for a549 cell. Viafect reagent provided optimal transfection efficiency and low toxicity for the majority of the cell lines tested, although there were some cell lines e. Achieve superior transfection efficiency while maintaining low toxicity lipofectamine 3000 reagent maintains a high transfection efficiency.
A comparison showing transfection efficiency of polyjet reagent vs. I have been transfecting a549 cells for long time but still could not get transfection efficiency more than 30%. Improved delivery of cas9 proteingrna complexes using. Cell plating a549 cells were seeded from 90100% confluent culture the day before the transfection. I found lipofectamine and geneporter2 might not only damage a549 also have worse tranfection efficiecy, so i used jetpei to transfection. To transfect cells with sirna, follow the protocol as described for dna but. Transfection protocol of adherent a549 cells 96well plate. Lipofection has become more common than the capo 4 precipitation, because it is just as simple to use but frequently provides a much better transfection efficiency. Singlecell transfection by electroporation using an. Comparative transfection efficiency of jetprime versus its main competitor. The reagent provides high transfection efficiency in many cells types and formats. I have used lipofectamine and fugene for plasmid transfection.
Transfection efficiencies were assessed by gfp signal. Xfect is a biodegradable transfection polymer, screened from a pool of 2,300 candidates for its very high transfection efficiency and low cytotoxicity profile. The commonlyused plasmid dna or rna lipofectamine transfection reagent had been shown to be able to deliver many proteins into cells, so we tested several lipofectamine variants for their ability to efficiently deliver cas9 rnps sells et al. In hacat cells, lipofectamine 2000 achieved a transfection efficiency. N v i s i on io 20 transit certified transfection reagents. Transfection protocol lipofectamine 3000 transfection. A reverse transfection protocol was used to transfect mcf7 and hepg2 cells. Transitx2 transfection reagent a dynamic transfection system for the. For additional information on scaling your transfection. It is no surprise that lipoplexes stick to the plastic surfaces of cell culture. Protocol for lipidmediated transient transfection in a549 epithelial. Ccl185 human lung carcinoma cells by fugene hd in 96 well plates.
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