I am planning to quantify the expression level of a receptor in certain brain region. The technical challenges for me are:
1. IHC or in situ: is it valid to use the normalized fluorescence intensity of the receptor to DAPI for quantification?
2. Of course, the more accurate methods are Western blot or qRT-PCR. However, my interested brain region is difficult to isolate for RNA extraction due to anatomical location and small area size. Is any good way to extract RNA from certain neuron types from a brain region?
Any advise or comment is welcome. Thank you!
Yes you can use IHC or FISH, although for IHC you would need to accurately dissect out the specific brain region and ensure that this is done consistently for all test groups for your comparison to be accurate (This would also hold true for FISH).Secondly you must ascertain the specificity and /or sensitivity of the receptor antibody you intend to use,and ascertain the characteristics of its staining in the brain.
This article might be useful
Cannabinoid CB2 receptors: Immunohistochemical localization in rat ...
cannakb.com/./Cannabinoid%20CB2%20receptors:%20Immunohistochemical%20lo...
by JP Gonga - 2005 -
Immunohistochemical study with measuring the optical density for the photographs is an easy method for the small hardly dissected areas in the brain. In my work; on doing qPCR for the hippocamus, nearly the same results as immunostain expression were obtained.
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