I have used different cell markers such as CD163 and CD206 for M2, but both can be expressed in M1. while for M1, I used 27E10 which can be expressed in M2 too.
1st, most macrophage have both M1 and M2 marker expression, so I do not think M1 M2 is a good concept. Just direct use the marker name that related with phagocytosis, antigen presentation...., it is much better than use the wrong concept M1 and M2
I think CD163 and CD206 are expressed in higher quantity in M2. The other markers you could use are IL-10 and Arg-1 (both highly expressed in M2 and barely detectable in M1). Conversely, M1 express higher levels of Il-12 and iNOS.
I recommend the following paper:
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3991396/
Article Resource Transcriptome-Based Network Analysis Reveals a Spec...
But, I could't get specific qualitative Ab against human M1 and M2 yet. particularly for immunofluorescent staining. Arg-1 more non-human macrophage related. CD163 and CD206 still expressed in M1 macrophages but in less quantitiy.
thanks for Simon and Michel for your PCR informations
1st, most macrophage have both M1 and M2 marker expression, so I do not think M1 M2 is a good concept. Just direct use the marker name that related with phagocytosis, antigen presentation...., it is much better than use the wrong concept M1 and M2
This is another good paper for answering your question: http://www.jimmunol.org/content/177/10/7303.full.pdf
We usually stained mouse mac intracellularly using anti-RELMalpha for M2/AAM phenotype. Staining was negative in non-helminth infected mice.
good luck
Completely agree with Dacheng Ding, M1 and M2 are too polar and require specific activation. Most macrophages express both, M1 and M2 markers.
Thanks for all of you, Mikhail I agree with you, but my problem is not the name.
my samples are monocytes isolated from PBMC and polarised with 20ng/ml IFN-gamma + 50ng/ml GM-CSF and other group polarised with 20ng/ml IL-4 + 50ng/ml M-CSF. I need to know if there any specific qualitative cell markers can be detected by immunoflourescent microscopy?
You can try intracellular staining for cytokines. But I think the best start is to do qPCR for iL12, il10, iNOS, Arg1 e.t.c
Hi Hassan,
MHC II, CD80, CD83, CD86 should have difference between your 2 groups if you do not use other factors i.e. TNF, LPS etc. Based on my experience, you could detect these marker expression level difference by confocal microscopy.
good luck
Dacheng
Right now, the M1 and M2 concepts are very questionable. Check out this article ( Immunity 2014, 41; 14-20), where you can find a figure (Fig 1) where the markers that corresponds to Macrophages / monocytes from mouse and human are summarized. Also, the article provide a clear view of the state of the M1-M2 concept.
Best Regards
Jorge Lloberas
Hi Joe,
As your request, here is my protocol.
1, setup reaction for a tagged gDNA lib.
- gDNA 10 ng to 500 ng
- 5X buffer 4 ul
- Enzyme-M 0.5 ul then add H2O to 20 ul
gently mix it throughly with pipet then incubate O/N in a PCR instrument
2, setup qPCR reaction:
purify and dilute the gDNA lib.
- diluted gDNA lib. 1 ul
- 2X PCR mix 10 ul
- primer 1 ul
- H2O 8 ul
3, set up thermal protocol depend on your PCR system and instrument.
4, calculate/quantitate based on CT value
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