Recently, I tried to differentiate THP1 into M2 macrophage using IL4 and IL13 (purchased from R&D). This is my protocol.
- THP1 was seeded into 6-well plate (10E6 cells/well). Incubate with PMA (100 nM) for 24 hours.
- After 24 hours, washing cells with PBS and incubate cells with fresh culture media (10% hea-inactivated FBS, 1% PS, RPMI) for 48 hours.
- Then, change the media and incubate cells with IL4 (20ng/mL) and IL13 (20 ng/mL).
- Harvest cells after 48 hours for qPCR (CD206 and Arg1).
I have a problem with qPCR that CD206 and Arg1 were not induced significantly... Please give me some advice...
Thank you.
Hi Long Le Hoang
Many thanks for your inquiry.
My suggestions would be to decrease the number of seeded cells and increase the time of PMA stimulation as per the attached protocol from the Bowdish lab.
In addition I would increase the duration of IL4 and IL13 stimulation prior to readout to 72 hours, as per Figure 5 from the attached paper.
Article M1 and M2 macrophages derived from THP-1 cells differentiall...
And for more detailed functional characterization, please consider:
Article Natural polymorphism of Ym1 regulates pneumonitis through al...
All the best & good luck with your experiments,
Michael
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