I have been using lentivirus, but am getting relatively low expression. Are there any specific medium requirements you recommend? Do you recommend serum? Also what do you recommend on medium changes?
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Improve lentiviral production using Lipofectamine 3000 reagent
Company: Thermo Fisher Scientific, Inc.
Job Title: R&D Scientist
We developed a Lentivirus production system using our Lipofectamine 3000 in our p-Lenti6.3/V5 (p-Lenti7.3/V5) Lentiviral Expression System. You can find the details of App Note as following website link: https://www.lifetechnologies.com/content/dam/LifeTech/global/life-sciences/CellCultureandTransfection/pdfs/Lipofectamine3000-LentiVirus-AppNote-Global-FHR.pdf
The LV packaging media is our specific media for Lentivirus production – Opti-MEM/GlutaMax with 5% FBS and 1X Sodium Pyruvate.
It is not necessary to change media post-transfection if you are planning to concentrate the lentiviral particles 24-48hrs post-transfection (with two supernatant harvests), or you can use benzonase treatment to eliminate the remaining DNA plasmids. However, if you are going to use crude lentivirus in your next step to transduce your target cells, the remaining DNA/reagent complex might be toxic to your cells, so changing the medium at post-infecting 6hrs is recommended.
Important to keep in mind, use crude lentiviral cell supernatant which had remained DNA/reagent complex to measure the viral titer using the %GFP method, there may be a slight increase in GFP positivity that could translate to a 1 fold higher titer. In this case, you would need to adjust your MOIs to account for the perceived increase when setting up your experiment.