Basic Immunohistochemistry

Use this category for questions regarding various immunological methods

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Basic Immunohistochemistry

Postby Lullabi88 » Jul 05 2016 10:34 am

Hi everyone,
I've just started my PhD in Neuroscience (mouse models of cognitive impairment) and unfortunately, I don't know much (yet! :) ) about tissue analysis.
As far as I've understood, there are three major techniques: immunohistochemistry, western blot and ELISA. I understand more or less how these techniques work and the different steps they involve, but I really don't understand how you interpret the results. Let's say I do immunohistochemistry on the hippocampus to identify expression of a certain receptor, how do I then quantify my results? How can I say that a certain group has more of these receptors than the other group? I guess that what I'm trying to say is, how do you go from the tissue images to the statistical analysis?
I have more or less the same question for the Western blot. What you obtain with the western blot is protein ladders. How do you read these ladders? Can you use Western blot to say whether a certain group has more expression of a protein than the other? Or does Western blot only tell you whether the protein is expressed or not? And again, how do you go from the ladders to the statistical analsis?

Thank you very much!!!!
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Re: Basic Immunohistochemistry

Postby relaxin » Jul 06 2016 9:06 pm

You can quantify fluorescent signal with sensitive imaging system with good software. But the equipment is very expensive and the quantification is time consuming.

For Western blot, it is acceptable to scan the intensity of the protein band of interest and get a semi-quantitative result. You need a scanner and an inexpensive software. It will not be "yes or no", but x-fold increase or decrease when compared with control. With three independent runs, you will get mean +/- SD.
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