I treat THP1 cells with 100 ng/ml PMA for 48 hrs and then stimulate them with 20 ng/ml of IL-4 and IL-13 for 48hrs (M2) and with 20 ng/ml IFN-gamma and 10 ng/ml LPS for 48hrs (M1). Then I leave them in fresh media for another 2-3 days. I use accuatse to detach them for FACs analysis. while CD11b confirmed monocyte to macrophage, I get negative population for CD206 in M2 macrophages and HLA-DR in M1 macrophages. I would appreciate if anyone can share experience with thp1 polarization and FACs. I do see evident morphological change, but thats not translating to my facs analysis.
Thank you.
Looking forward to hearing suggestions.
Hi,
the M2 polarization of THP1 is very difficult to obtain, we never obtained a CD206 positive population with an IL10 IL4 stimulation nor with dexamethasone, only a moderate positivity for CD163 and CD209.
For M1 markers you can try CD86 , it worked pretty well with an IFN/LPS stimulation
good luck
Sébastien Banzet Do you mind sharing your protocol for IFN/LPS stimulation?
Hi @Meenakshi Sudhakaran
I have issue with the polarization of THP-1 cells into M2 type too. They do not express CD206. On the other hand, THP-1 cells treated with PMA, IFNg and LPS clearly expresses HLA-DR. THP-1 cells were primed with 200 ng/mL PMA for 6h, and added 20 ng/mL IFNg and 100 ng/mL LPS followed by incubation for futher 66 h. Medium was chaged to fresh one and incubated for 48 h.
Taichi Matsumoto Hi. Did you get a positive population of HLA-DR for M0 macrophage? (PMA 200 ng/ml for 72 hrs)?
Meenakshi Sudhakaran No, HLA-DR was negative on THP-1 cells treated with 200 ng/mL PMA for 72h.
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