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GFP expression has been observed for up to one week as measured by light and fluorescence microscopy.
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The pmaxGFP™ Vector provided in our Nucleofector® Kits is not expressed in insect cells. We strongly recommend an insect expression vector encoding a fluorescent protein or lacZ reporter as a positive control for your experiments [e.g. Novagen™’s...
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Insect cells are cultivated at lower temperatures than E.coli or mammalian cells (S2 cells at 24°C, Sf9 cells at 28°C).Therefore they need a dedicated incubator with appropriate temperature.
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It is unlikely that plasmid size would affect in vitro differentiation capability. Gene targeting constructs are often as large as 20 kb and if treated well the ES cells are perfectly capable of generating germ line chimaeras afterwards (indicating...
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Vectors with a CMV promoter typically work fine for transient protein expression in mammalian cells, like HEK293, CHO or NSO. For insect cells, like Sf9 or Sf21, vectors with insect promoters are required. In HEK293E (EBNA) cells, the use of...
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The optimal incubation time ranges from 2 to 6 days depending on various factors like the stability of the protein in the medium, potential toxicity of the protein, cell density and stability of the plasmid in the nucleus. To check the timepoint for...
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In general, all reporter genes can be used.For some applications (e.g. reporter gene experiments in various primary cells, experiments with late analysis time point due to starving and /or stimulation) a reporter gene with a longer half life than...
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Yes. Stimulate the cells with anti CD3 and anti CD28 antibody for 24 hours. Use the optimized protocol. Often times, this will result in higher transfection efficiency (60-70% is possible). Alternatively, you can stimulate with ConA and IL-2 for 2-3...
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This totally depends on the Nucleofector® system you are working with. While in the 100 µL reaction vessels usually 1 - 5 µg DNA is being used, for the 20 µL reaction vessels (e.g. X Unit, 96-well Unit and HT...
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For many cell lines in suspension, especially sCHO, conventional transfection methods are very limited in efficiency. For protein production, suspension cells are favored over adherent cells because they are much easier to cultivate. Unlike...
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