Immunohistochemistry: What am I looking at? - (Jan/20/2012 )
This is a very general and vague question but I really don't know any other way to ask: I've just started working in a lab and began IHC. I section frozen tissue samples and then proceed with the IHC protocol. I produce an image.
But I really don't quite understand what I'm looking at. I understand how IHC works- the binding of the primary to the desired protein and then the binding of the secondary tagged with a fluorphore to the primary, etc. But I suppose I don't recognize the tissue behind the staining? Is IHC mainly used to determine the presence or absence of a protein of interest? I can do that, although I wouldn't be able to tell where exactly the protein is. Can you glean any more information from IHC?
I'm using sections from a mouse. I've looked up some pictures of what tissue sections from them are supposed to look like, but, after staining, the tissue looks nothing like them. How do people read these things? Any advice or helpful websites, books, etc would be greatly appreciated.
Yes, you can obtain information regarding the presence or absence of a protein, its expression level, its location in (nuclear or cytoplasm) or outside the cell.
To be able to view IHC slides, I assume you have done counterstaining which makes cellular structuer clear to see, you need to have some knownledge about the organ or tissue you are working with. This is usually taught in histology class. There are many online resources providing images of mouse or human HE stained tissue sections, such as this one The Visible Mouse Project at http://tvmouse.compmed.ucdavis.edu/
Thanks for your reply pcrman. Yes, I use hoescht to counter stain. I've never taken histology so perhaps that is my problem. I'll try to self-educate myself on the topic. Thanks again.