Do you know if your primary cells can grow longer in culture without any virus infection? What is the maximum length of growth you have seen in your primary cells after differentiation?

The best way to introduce your lentivirus could be to use an established cell line (if available/or applicable) instead of primary cells.

Hello Felix,

I would recommend to differentiate your cells first

and after differentiation you can try to overexpress the gene of your interest.

If the lentiviral transduction does not work, you can try to transfect your differentiated dendritic cells with mRNA. We have established a protocol that allows highly efficient transfection of primary murine bone marrow-derived macrophages (BMDM) with in vitro transcribed mRNA. This protocol should also work with dendritic cells.

If you are interested, you can have a look at our very detailed protocol for differentiation and transfection of BMDM:

https://www.researchgate.net/publication/337153549_Highly_Efficient_Transfection_of_Primary_Macrophages_with_In_Vitro_Transcribed_mRNA

Kind regards,

Alex

It depents on what type of primary immune cells you are handling with and the purpose of the preparation.

If it is T cells or NK cells, lentivirus would result high infection efficiency and you can later use them for both in vitro and in vivo eperiements.

If you are working with cells that are resistent to lentivirus such as macrophages, then adenovirus of specific serotype would help you. As its has been reported Ad5f35 adenovirus infect macrophages at a high efficacy.

Good luck!