We are using iba1 as the marker for the activation and proliferation study of microglia following focal cerebral ischemia. However iba1 stains both macrophages and microglia thus creating confusion in immunohistochemistry. Could you please let me know the best marker that only stains microglia following stroke challenge ?
There is no specific marker in immunohistochemistry that only stains microglia and not macrophages. All antibodies stain macrophages and microglia, especially activated microglia. You can differentiate resting or partially activated microglia based on morphology vs macrophages but once microglia become phagocytic this is not possible. Usually resting microglia express lower levels of many markers with an increase upon activation.
This is an area that the field has struggled with for decades and has hindered research into the role of microglia as compared to macrophages. ED-1 and CD11b (see the antibody product sheets) are expressed on macrophages, though much less on resting microglia.
Hi Bhakta
I suggest you consider using CD11b.
I am not sure what exactly are you trying to stain and what is your procedure, but CD11b might be a proper marker for you.
Boaz
Hello..You can try ED-1 which is a specific marker for only microglial activation. It will eliminate the resting and intermediate stage microglia and only show activated stage microglia. It may help you.
Gud luck..
There is no specific marker in immunohistochemistry that only stains microglia and not macrophages. All antibodies stain macrophages and microglia, especially activated microglia. You can differentiate resting or partially activated microglia based on morphology vs macrophages but once microglia become phagocytic this is not possible. Usually resting microglia express lower levels of many markers with an increase upon activation.
This is an area that the field has struggled with for decades and has hindered research into the role of microglia as compared to macrophages. ED-1 and CD11b (see the antibody product sheets) are expressed on macrophages, though much less on resting microglia.
I don't have it at hand, but look into Kreutzberg's publications on microglia mostly related to the facial nucleus. However Timothy Seabrook made the point.
Timothy Seabrook is right, on all his points. All commercially available markers of microglia in the uninjured brain (Iba1, CD11b F4/50, CX3CR1 etc) are present on all/at least subsets of infiltrating monocyte derived macrophages in the injured brain.
Butovsky et al Nat Neurosci 2014 propose a microglia specific expression signature, which includes P2ry12 and FCRLS, however, the former is down regulated during some inflammatory scenarios and neither are commercially available.
Differential CD45 expression measured with flow cytometry can give you quite a good estimate of the two populations, though CD45 is thought to be highly expressed in very activated microglia, so confusing a small subset of them with infiltrating macrophages.
Hi! Timothy and Andrew have given good suggestions. I am using flow cytometry to distinguish microglia and monocytes and it is impossible to differentiate them via CD45 and CD11b in inflammatory conditions as microglia also become CD45 and CD11b high. Best is to perform CD45.1 bone marrow chimera to distinguish between them in flow cytometry.
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