Transient plasmid dna transfection into hela and hacat. Refer to the cell lines database for a list of cell types successfully. Transfection of nih3t3 cells, hela, swis 3t3, 293t with. However, few studies have examined cellular responses to this delivery system. Lipofectamine 2000 cd reagent is a proprietary animal originfree formulation for transfecting. Help frozen lipofectamine transfection and transduction. It consistently produces high transfection efficiency and high protein overexpression.
I use lipofectamine 2000 invitrogen for transfection of both sirna and plasmid dna into these 293 cell lines. Versatile transfection reagent that has been shown to effectively transfect the widest variety of adherent and suspension cell lines. Culture cells in serumfree media that are free of animal. Works effectively with all common cell lines as well as with many challenging ones, and can. Test serumfree media for compatibility with lipofectamine 2000 since some. Lipofectamine contains lipid subunits that can form liposomes in an aqueous environment, which entrap the. Invitrogen lipofectamine 2000 transfection reagent is a versatile transfection reagent that has been shown to effectively transfect a variety of payloads into the widest variety of adherent. The instructions are recommending that you do this in duplicate with the two indicated different concentrations of lipofectamine 3000. To determine the required amount of lipofectamine 2000 for forming of lipoplexes, different amounts of lipofectamine 2000, 1, 2, 4, 6. Invitrogen lipofectamine 3000 transfection reagent. Lipofectamine 2000 transfection reagent from invitrogen. We tend to follow the protocol provided, which is simple and easy to follow. Optimizing transfection lipofectamine 2000 cd reagent.
Lipofectamine 2000 reagent is a lipidderived reagent designed to. The trusted, simple, broadspectrum transfection reagent for most cell lines. Lipofectamine 2000 reagent thermo fisher scientific. The intracellular trafficking mechanism of lipofectamine. Despite this, a satisfactory mechanismbased explanation of their. I use 24well plates for transfections in triplicates. When we add the fullserum media back 6 hours later, we use antibioticfree media. Invitrogen lipofectamine 2000 transfection reagent 1. Transfection of nih3t3 cells, hela, swis 3t3, 293t with lipofectamine 2000 1. Researchers use lipofectamine 2000 reagent for sirna and shrnabased gene knockdown experiments, as well as for gene expression studies. Test serumfree media formulations for compatibility with lipofectaminez 2000 as some serumfree formulations can inhibit cationic lipidmediated transfection.
Theres a difference between the two protocols lipofectamine 3000 has an extra reagent that needs to be added. Help frozen lipofectamine aug092006 hi i am really worried, i transported my lipofectamine from one lab to another in a dry ice container, and when i arrived to my lab and i open the box i found that the lipofectamine has been frozen i put it immediatly at 4 c, but i dont know if i damaged the lipofectamine can i still use this. Lipofectamine 2000 is then added to culture medium within a sterile tube along with the dna at a 3. Genscript recommends using lipofectamine 2000 for all transfections. Cd 293, 293 sfm ii, and vpsfm are media formulations known to inhibit. Rna lipofectamine rnaimax complexes must be made in serum free medium such as optimem. What is the appropriate way of adding lipofectamine 3000 to cells. Highest transfection efficiency in many cell types and formats e. Transfect plasmid dna and sirna at the same time using lipofectamine 2000 reagent by adding 30 pmol 0. Lipofectamine 2000 transfection reagent is a versatile transfection reagent that has been shown to effectively transfect the widest variety of adherent and suspension cell lines. Lipofectamine 2000 from invitrogen biocompare product. Invitrogen lipofectamine 2000 transfection reagent is a proprietary formulation for the transfection of nucleic acids dna and rna into eukaryotic cells providing the following advantages. Dnaplus lipid complexes must be made in serumfree medium such. Descriptionlipofectamine 2000 is a proprietary formulation suitable for the transfection of.
A hela cells were transfected with hcv dsrna using lipofectamine 2000 reagent. Lipofectamine 2000 is a lipid based transfection reagent for eukaryotic cells. A lot of the transfection protocolefficiency depends on the kinds of cells you are using. Lipofectamine 3000 reagent is a proprietary formulation for transfecting nucleic acids into a wide range of eukaryotic cells and especially designed for hard to transfect cells. Invitrogen lipofectamine 2000 transfection reagent. Lipofectamine 2000 transfection reagent from thermo fisher. The microscopic analysis was performed using antirigi mab. Life technologies corporation andor its affiliates warrant their.
Lipofectamine 2000 transfection for hek293 cells about 6070% transfection rate transfect cells 6well plate. Lipofectamine 3000 protocol written out with timing. The purpose of this study is to evaluate the effect of sirna transfection using lipofectamine 2000 on cellular autophagy. 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. You can generally use other serumfree medium instead of optimem. Lipofectamine 2000 dna transfection reagent protocol. Dilute lipofectamine3000 reagent in optimem medium 2 tubes, mix well follow the instructions as indicated.
Transfections were performed in 24well plates using 0. Lipofectamine or lipofectamine 2000 is a common transfection reagent, produced and sold by invitrogen, used in molecular and cellular biology. Optimized conditions for successful transfection of human. I tried reversetransfecting hepg2 cells using lipofectamine 2000. Lipofectamine 2000 and lipofectamine rnaimax transfection. Invitrogen lipofectamine 2000 transfection reagent is a proprietary formulation. Each reaction mix is sufficient for triplicate 96well, duplicate 24well, and single well 6well transfections, and accounts for pipetting variations. Dmem invitrogen 11965092 with 10% fbs with no antibiotics 4. Effects of lipofectamine 2000sirna complexes on autophagy. Transfection commonly refers to the introduction of nucleic acids into eukaryotic cells, or more specifically, into animal cells. Lipofectamine reagents are widely accepted as goldstandard for the safe delivery of exogenous dna or rna into cells. Although invitrogen recommends their own brand of culture medium for transfection, called optimem, we found no difference between this and conventional media.
Lipofectamine or lipofectamine 2000 is a common transfection reagent, produced and sold by invitrogen. Lipofectamine 3000 provided gentle but efficient transfection in several line of interest to our lab, heioc1 mouse cochlear 69fold, nccit and ntera2 3 fold testicular cancer lines and allowed for very nice dose response assays for cisplatin sensitvity assays. For researchers interested in other delivery methods, we also offer additional invitrogen lipofectamine. Lipofectamine rnaimax has a broad peak of activity. Application note gene delivery solutions stem cell. The animal origin free formulation ensures that mammalian cell culture and bioproduction processes are free of animalderived materials. Invitrogen lipofectamine 3000 transfection reagent lipofectamine 3000 transfection reagent leverages our most advanced lipid nanoparticle technology to provide superior transfection performance with improved application outcomes and reproducible results. Make dna lipofectamine 3000 complexes in serum free medium such as. Product isguaranteed for 6 months from the date of shipment if stored properly. Make dnalipofectamine 3000 complexes in serumfree medium such as. Lipofectamine rnaimax transfection reagent is a proprietary formulation for transfecting small rnas e. It is used to increase the transfection efficiency of rna including mrna and sirna or plasmid dna into in vitro cell cultures by lipofection.
Invitrogen 11668019 view more versions of this product. Lipofectamine 2000 transfection reagent dartmouth college. For lipofectamine plus transfections, the dna was preincubated with 4. Photograph of expected results dna lipofectamine scaling up or down. Lipofectamine 2000 transfection reagent is versatile transfection reagent that has been shown to effectively transfect widest variety of adherent and suspension cell lines. Mix gently and incubate for 5 min at room temperature. The lipofectamine 3000 is the bees knees though, i managed to get loads of transfects from a difficult to transfect cell line that had been giving us trouble with the lipofectamine 2000. The optimal level of each transfection reagent was determined empirically and all reagents were used according to manufacturers protocol. Lipofectamine 2000 is commonly used for sirna transfections.
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