I would like to analyze murine bone marrow derived macrophages (myeloid) and I am looking for the most accurate staining panel for flow cytometry.
It does not need to be specific for pro- and anti/inflammatory macrophages.
Are CD11b and F4/80 sufficient?
Is it necessary to include Ly6C and/or Ly6G?
Happy to hear your opinions.
Thank you.
M-CSF-generated BMDMs should be 99.9% pure by day 7. CD11b and F4/80 are good markers, I'd also use CD64 as a macrophage marker and LyG and Siglec F, to ensure absence of neutrophils and eosinophils, especially if working with BMDMs for the first time.
Hi, as Alexander says, I would recommend using CD11b instead of F4/80. It tends to work way better. Also I would include CD11c for dendritic cells which should be negative. Normally after a 6-7 days of BMDM culture I obtain a 1% CD11c+. That would tell you the purity of your culture, for a more specific panel CD28, CD206 and Ly6G and/or GR1 would be good as well.
Thank you for your answers.
For this particular screen, I won't be generating BMDM, it is a basic screen for macrophages from freshly isolated bone marrow.
Since I expect some cells to express GFP, I am basically screening for the GFP positive population in order to generate BMDMs from GFP positive cells only in a second step.
For now, just screening for GFP in the various populations in total BM.
Ah, alright. I have done a full screening of the Bone Marrow just flushed out from the mouse tibia for the following panel: CD3, CD45R (B220), F4/80, CD11b, Gr1 (Ly6G) and CD11c. I can tell, you will have a massive population of CD11b and then High GR1 and moderate F4/80. CD45R it's way lower and CD3 almost nothing.
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