We have had success using lipofectamine reagents in the past and the lipofectamine 3000 appears to be the best option. Transfection of nih3t3 cells, hela, swis 3t3, 293t with. Add the oligomerlipofectamine 2000 complexes to each well containing cells and medium. Not recommended for use with lipofectamine 2000 or lipofectamine rnaimax. Lipofectamine 3000 is a transfection reagent that was developed to improve transfection efficiency in many difficult to.
Mix gently and incubate for 20 minutes at room temperature solution may appear cloudy. Dilute 4 amounts of lipofectamine 2000 reagent in optimem medium. For additional information on scaling your transfection reaction, see page 4. Mix optimem and lipofectamine in another eppendorf 32 l lipofectamine in 0,8 ml optimem 3. Lipofectamine 3000 reagent outperforms invitrogen lipofectamine 2000 and fugene hd reagents. In this study, we demonstrated that using an improved transfection reagent, such as lipofectamine 3000, helps to eliminated many of the pain points. Prewarm 50ml of optimum stored in cold room at 4c 10min. Highest transfection efficiency in many cell types and formats. Invitrogen lipofectamine 2000 transfection reagent 1. Lipofectamine or lipofectamine 2000 is a common transfection reagent, produced and sold by invitrogen, used in molecular and cellular biology. Mix optimem with dna in an eppendorf 510 g dna0,8 ml optimem 2. Transfect plasmid dna and sirna at the same time using lipofectamine 2000 reagent by adding 30 pmol 0. Transfection protocol lipofectamine 3000 transfection reagent. Also, my molecule of interest is an antisense oligonucleotide.
Lipofectamine 2000 and 3000 summary in this comparative study, transfections were performed with dnain and compared to the current market leader, lipofectamine 2000 lf2k, using a series of cultured celllines and primary cells. 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. The product manual includes advice on how to optimize transfection efficiency, as a number of conditions can be varied including dna. Dnaplus lipid complexes must be made in serumfree medium such. Add 100l of optimum to the eppendorf tube to dilute the dna and mix by tapping. You can search for similar products in our product directory. How does transfection with lipofectamine 2000 work. Highest transfection efficiency in many cell types and formats e. Lipofectamine 2000 transfection reagent is versatile transfection reagent that has been shown to effectively transfect widest variety of adherent and suspension cell lines. Use a range of several ul of lipofectamine and ug dna to increase viability after transfection and efficiency. Each reaction mix is sufficient for triplicate 96well, duplicate 24well, and single well 6well transfections, and accounts for pipetting variations.
Life technologies corporation andor its affiliates warrant their. Step tube complexation components amount per well 24well 1 tube 1 optimem i medium 25 l lipofectamine 3000 reagent 1. Lipofectamine 2000 transfection reagent from invitrogen. Add 6l of lipofectamine2000 from biostores and 100 l of. Lipofectamine 3000 reagent thermo fisher scientific fr. Lipofectamine 2000 transfection reagent is a proprietary formulation. Versatile transfection reagent that has been shown to effectively transfect the widest variety of adherent and suspension cell lines. Invitrogen lipofectamine 3000 transfection reagent from.
Prior to mixing with dna, lipofectamine 2000 should be diluted in optimem i. Lipofectamine 2000 reagent dna seed cells so they are 7090% con. Formulation of lipofectamine 2000 and lipofectin signagen blog. Lipofectamine 2000 reagent thermo fisher scientific. Biomodule transfection and control user manual unit with. Lipofectamine 2000 is then added to culture medium within a sterile tube along with the dna at a 3. Fibroblast 3t3, cos7 myoblast c2c12, l6 crl1458 hepatocyte hepg2, huh7 erythroleukemic cell k562. Lipofectamine 2000 dna transfection reagent protocol. Make sufficient 10% fbs in optimem media wo antibiotics sterile filter 2.
Lipofectamine plasmid transfection bridges lab protocols. 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. Invitrogen lipofectamine 2000 transfection reagent life. I use 24well plates for transfections in triplicates. The vector contained an ofp reporter gene and was transfected with lipofectamine 2000 or lipofectamine 3000 reagent into a u2os and b hepg2 cell lines. Lipofectamine 2000 is a highperformance transfection reagent for gene expression and gene silencing. For more information about lipofectamine 2000 and other transfection reagents, refer to our web site or contact technical support page. Theres a difference between the two protocols lipofectamine 3000 has an extra reagent that needs to be added. Transfection of nih3t3 cells, hela, swis 3t3, 293t with lipofectamine 2000 1. Lipofectamine 2000 transfection reagent dartmouth college. Invitrogen 11668019 view more versions of this product. Cells in a 6 well dish, plated and at 90% confluence. Whats the difference between lipofectine and lipofectamine. Transfection of mcf7 with lipofectamine 2000 maartje vogel.
What is nice about invitrogens lipofectamine 2000 is that it has an extensive, very useful manual which addresses these issues and helps. Lipofectamine 2000 cd chemically defined transfection reagent is a proprietary animal originfree formulation for transfecting nucleic acids into eukaryotic cells. Lipofectamine 2000 reagent delivers dna or sirna with excellent transfection performance for protein expression, gene silencing, and functional assays. Invitrogen lipofectamine 3000 invitrogen lipofectamine 3000 transfection reagent 1. Transfection commonly refers to the introduction of nucleic acids into eukaryotic cells, or more specifically, into animal cells. Depending on the size of culture vessel being used, the lipofectamine 2000 is diluted between 25 and 50fold.
Product is guaranteed for 6 months from the date of shipment if stored properly. Plasmid dna, synthetic sirna, rnai plasmids shrna, mir content and storage. The average lentiviral titers in our preparations are around 56x 10 5 x 10 7. Invitrogen lipofectamine 3000 transfection reagent. Transfection protocol lipofectamine 3000 transfection. Researchers use lipofectamine 2000 reagent for sirna and shrnabased gene knockdown experiments, as well as for gene expression studies. Each reagent was used to transfect hek 293, hela, lncap, hepg2, and a549 cell lines in a 96well format, and gfp expression was analyzed 48 hours posttransfection. Lipofectamine 3000 is a transfection reagent that was developed to improve transfection efficiency in many difficult to transfect cell types. Contact life technologies technical service or your local sales representative for permission to crossreference the dmf.
Lipofectamine 2000 reagent protocol thermo fisher scientific. Add diluted dna to each tube of diluted lipofectamine 2000 reagent 1. Life technologies corporation andor its affiliates warrant their products as set. Mix each tube well by vortexing or flicking the tube. Lipofectamine 2000 reagent protocol 202lipofectamine 2000 dna transfection reagent protocol transfect cells according to the following chart. Invitrogen lipofectamine 2000 transfection reagent. Optimizing transfection lipofectamine 2000 cd reagent. The virapower lentiviral support kits includes the virapower packaging mix, lipofectamine 2000, and a selection agent. Advanced transfection with lipofectamine 2000 reagent. 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. Lipofectamine 3000 reagent maintains a high transfection efficiency within a robust dynamic range of lipid doses for quick and easy optimization.
At the foundation of the popularity of this broadspectrum reagent is the reliably high efficiency and simple protocol for success across a broad range of cell lines figure 1. Lipofectamine transfection of c2c12 cells bridges lab protocols. The transfection reagent lipofectamine 2000 lf2000, invitrogen is preferred for transfection. Aug 07, 2012 add 2 microliters of lipofectamine ltx reagent to tube 1, 3 microliters to tube 2, 4 microliters to tube 3 and 5 microliters to tube 4. I personally have used this reagent to transfect cells for fluorescent microscopy, gst pull downs, and coimmunoprecipitation experiments. 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. Lipofectamine 3000 reagent is designed to efficiently transfect difficulttotransfect cells, yielding superior transfection performance across the broadest array of cell types. 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.
The trusted, simple, broadspectrum transfection reagent for most cell lines. Types of kits this manual is supplied with the kits listed below. I use lipofectamine 2000 invitrogen for transfection of both sirna and plasmid dna into these 293 cell lines. Invitrogen lipofectamine 2000 transfection reagent is a proprietary. Works effectively with all common cell lines as well as with many challenging ones, and can. Lipofectamine 2000 transfection reagent from thermo fisher. Although invitrogen recommends their own brand of culture medium for transfection, called optimem, we found no difference between this and conventional media.
The virapower and virapower ii lentiviral expression kits include the virapower lentiviral support kit plus an expression vector and the. Effi ciency and ofp expression were analyzed 72 hours posttransfection and a u2os and b hepg2 cells showed 4fold and 80fold improvement with lipofectamine. The cells were trypsinized after 48 hours, resuspended in. Transfections were performed in 24well plates using 0. Using lipofectamine 2000 cd reagent for transfection provides the following advantages. Lipofectamine 2000 cd reagent is a proprietary animal originfree. Lipofectamine 2000 transfection protocol 10cm plates reagents. Photograph of expected results dna lipofectamine scaling up or down.
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