Hello,
I am having trouble with transient transduction of lentivirus particles in THP-1 cells and differentiated macrophages. I do not have an antibiotic resistance gene in my vector to do a selection. Instead, I have a reporter gene readout (tdTomato). My transduction efficiency is really low (less than 10%) and I have added polybrene and done spinoculation (1000g for 45 mins at 37˚C). I have even tried transducing the cells twice but still no luck. I tried scaling down from 6 well plate to 12 well and still don't have overexpression. I have read about the SAMHD1 block but I tried with macrophages and thp1 (thp-1 do not have the block apparently).
I am unsure what to do next because I know that the issue is not from my lentivirus (they work fine with other cell types). Any tips or things I can try to get results? Have any one of you had experience with transient transduction of monocytes?
I had similar issue with THP1 transduction, also had low efficiency.
What I did to improve:
1. Double transduction
2. FACS sort GFP positive in your case Tomato and then expand those positive population.
Other Suggestions
-Check titters of your LVs
- Play with LVs vs cell ratio, I would suggest to go bit higher ratio
- Do transduction 10 cm dish, so that you have enough cells for FACS sort
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