**1. Introduction**

Immunohistochemistry (IHC) represents a way to build a picture of particular distribution and localization of molecular markers within cells and in the proper tissue context and is a powerful tool that provides important diagnostic, prognostic, and predictive information supplemental to the morphological assessment of the tissues. Although less sensitive quantitatively than such immunoassays as western blotting or ELISA, IHC enables observation of molecular signature in the context of intact tissue. In its very simplified method, IHC visualizes target antigens by using target-specific antibodies tagged with appropriate labels. However, lack of need for labeling of molecular marker-specific primary antibodies and higher sensitivity made indirect staining methods as the preferred staining method. The need for more sensitive detection systems in case of minimally expressed markers was a provocative factor that eventually led to the emergence of next generations of IHC detection methods with the hope to amplify staining signal. IHC

methods based on avidin-biotin interaction and polymer- and tyramide-based signal amplification are among IHC signal amplification methods that have greatly enhanced the sensitivity of IHC staining. However, when more sensitive methods are used, background signal tends to increase along with the target signal and so highly sensitive detection systems are not always desirable. Therefore, the optimal IHC method is planned as a compromise between sensitivity that allows proper and reliable visualization of a given molecular marker and at the same time avoiding background signals that impair staining index and specificity of the staining method. In an optimal IHC detection system, tissue type, level of expression of the marker of interest, localization of the marker, and cost are among important factors that should be taken into consideration. As a general rule of thumb, there is no a *bona fide* IHC detection method that is universally accepted. Although it does not rely on chemical reactions that take place in IHC, immunofluorescence (IF) staining follows almost the same rules as with IHC and so concerns on detection systems that are also applicable to IF staining methods. In this chapter, we will focus on detection methods in immunohistochemistry and immunofluorescence stainings and highlight in detail potential application, advantages, and disadvantages of each method.
