Hi,
I am transducing jurkat cells with lenti-virus but my efficiency rate is low... my transducing method is this: 1. I defreez viruses and put them 20 min in room temp. , 2. resuspending them in PBS and pipette it well. 3. 100.000 cell /96-well + proper concentration of viruses and pipetting again. ( I tested different concentrations) 4. incubation for 2 h. 5. transfer them to 24-well plate.
6. incubate 7. observe my cells after 72 h.(even up to 5 days) by fluorescent microscope. (mcherry).
I see very pale red color but about 80-90% of cells are red!!
NOW, how can I enhance the transduction rate to have sharp red?
You can increase efficiency of transduction using: 1) Polybrene (a cationic polymer that can greatly enhance the efficiency of the retroviral or lentiviral infection to the mammalian cells), 2) spinfection (centrifugation of the cells together with virus - increases contact between tumor cells and the virus) or 3) consecutive transduction (doing it twice on the same group of cells). Polybrene gives the best results, but does not work great with all cell types.
However, if the cells after transduction should be red, and you have 80-90% of them expressing mCherry, then the efficiency of transduction is great. I only got better efficiency when I was transducing HEK293 cells, and they are transduction control and very, very easy to transduce.
I would recommend review/optimize your thawing step and conditions, there is a possibility you compromise the integrity of your lentiviral vector, also make sure your cells show a great viability (at least 80%). Jurkat cells are T cells and should be suspending not adherent making transduction a bit more tricky than adherent ones like HEK293. If you are planning to make cells stably expressing whatever your gene, you can use polybrene as Patrycja mentioned (usually at 8 ug/ml of your culture medium)
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