Immunohistochemistry is a method for demonstrating the presence and location of proteins in tissue sections. Though, less sensitive quantitatively than immunoassays such as Western blotting or enzyme linked immune sorbent assay (ELISA), it allows the study of the distribution and localization of specific cellular components by providing supplemental information to the routine morphological assessment of tissues. This is especially useful for assessing the cellular markers that define specific phenotypes then providing important diagnostic, prognostic, and predictive information relative to disease status and biology. In general, the information gained from IHC combined with microscopy literally provides a “big picture” that can help make sense of data obtained using other methods.
I would always start with IHC and move toward WB. Combined together they are powerful findings. Also, sometimes could be easier to obtain and quantify IHC data vs. WB. It depends from your time, resources and preference.
There are numerous IHC methods that may be used to localize antigens, such as: direct and two step indirect method. The selection of a suitable method should be based on parameters such as the type of specimen under investigation and the degree of sensitivity required. Immunohistochemistry involves specific antigen-antibody reaction; it has apparent advantage over traditionally used special enzyme staining techniques that identifies only a limited number of proteins, enzymes and tissue structures. Therefore, it has become a crucial technique and widely used in many medical research laboratories as well as clinical diagnostics.
I would always start with IHC and move toward WB. Combined together they are powerful findings. Also, sometimes could be easier to obtain and quantify IHC data vs. WB. It depends from your time, resources and preference.
Immunohistochemistry confirms protein expression of selective genes in a sense that, this technique is looking at proteins. A higher gene expression in a given situation does not automatically mean that the protein is also highly expressed.
Both western blot and IHC can be used to partially validate the information given by the other technics. Both of those technics could be considered as semi quantitative as they have both potential limitations (i.e optical limitation, generated by saturation level of pixels, number of steps, amount of protein used..).
Those limitations can be solved by technical adjustments.
Once those are fixed, IHC (and WB) can be used on purpose of validating gene expression
The IHC has also the advantage of providing information on the cellular and the sub-cellular localizations of the protein of interest.
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In general, the hypothesis that gene over-expression leads to protein over-expression has a very weak support by experimental evidence. The truth is hidden. We see that no correlation is much more likely. I know only one notable exception: Her2 gene amplification in breast cancer correlates well with protein overexpression which we routinely detect my IHC and use a surrogate test for gene amplification status because it is easier and cheaper to do than FISH/CISh or genemicroarrays/real time PRC etc. I know abundant examples that the same is not happening for many other targets. Therefore such interaction should be target specific and overgeneralization will not work for "gene overexpression-protein overexpression dogma". These are just my thoughts and I agree with the other posts.
Proteins are molecules that are the final product of information provided by genes. Proteins "work" in the cell, while mRNA is only a vehicle that transfers information from the gene. The whole exciting RNA interference system is able to silence gene expression at the mRNA level and therefore you can never be sure whether protein expression correlated with the initial DNA expression at the mRNA level! I was always most happy with the protein expression, and if it is also localized to a specific cell of a tissue- that is very informative. E.g. retinoblastoma protein was thought to be ubiquituous because of its central role in the cell cycle control (assessed with the Western blot). However, we found that its expression depends on the degree of cell maturity!
L Szekely, WQ Jiang, F Bulic-Jakus, A Rosen, N Ringertz, G Klein and KG Wiman. Cell type and differentiation dependent heterogeneity in retinoblastoma protein expression in SCID mouse fetuses. Cell Growth & Differentiation, Vol 3, Issue 3 149-156, 1992
You can please refer to the links below to obtain certain crucial informations:-
1. http://cancerres.aacrjournals.org/content/55/24/6006.short
2. http://www.sciencedirect.com/science/article/pii/S0740257015000179
3. http://www.ncbi.nlm.nih.gov/pubmed/24164438
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