Hey Gang,
Very easy question to someone who knows more about identifying glia than I do! I am using antibodies against S100 to mark astrocytes and Iba1 to mark microglia in a culture of primary cortical neurons (from E17 mouse, DIV7). It seems that S100 labels not only what appear to be astrocytes, but also the nuclei of neurons, and some very interesting looking cells that I would guess are microglia (attached). Does anyone have experience with S100 for IF, does it detect many young neural cell types broadly? Do these look like microglia to you? The cells positive for Iba1 look nothing like these (attached), so I'm wondering if it's better to make a determination based on morphology in case my Iba1 antibody is not so good and S100 is too broad. Thanks for your input!
Hi Sara,
Just had a quick look at your images, and you right that there seem to be a mismatch between the morphology and immunolabeling of the cells. I would suggest that you try your antibodies in cortical slices from E17 mice and compare the immunolabelling and morphology. This should help you clarify if the culturing conditions are causing an abnormality?
Best wishes,
Refik
Hi Sara,
I haven't used S100 in this context, but it is fairly well known in a diagnostic IHC context that S100 is really not that specific. If you want to identify astrocytes you should shoot for a more specific antigen, like GFAP for instance. IBA1 should be pretty specific for microglia.
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