Immunohistochemical detection of metallothionein

Zinc (Zn (II)) is an essential microelement, it has critical role in normal health and development [1]. Binding of Zn (II) to zinc coordination motifs in proteins stabilizes the structure or influences the function. The prevalence of genes encoding zinc proteins is estimated to be over 3% of the 32,000 identified genes. Over 300 Zn (II)-dependent enzymes have been defined and characterized [2]. It has been shown that Zn (II) can regulate the DNA-binding activity of zinc finger transcription factors [3]. The Zn (II)-metallothionein (MT)/thionein pair, which is an important component of cellular Zn (II) homeostasis, is critical to sequester or release Zn(II) depending on the local redox state, thereby influencing the function of numerous enzymes and transcription factors that control cell proliferation, apoptosis and signalling pathways [4,5]. Abnormal MT function and expression have been implicated in various human diseases, including cancer [6]. There are at least 10 isoforms of MT in human body, which are expressed in a tissue specific pattern and may play distinct roles in the various cell types. MT-I and MT-II isoforms are present in all cells throughout the body, MT-III was first isolated as a growth inhibiting factor (GIF) from brain neurons, MT-IV is located in stratified epithelium [6]. Transcription of MT-I and MT-II can be induced by inflammatory cytokines (IL-6, TNF-α, interferons), lipopolysaccharids, glucocorticoids, free radicals, antioxidants or heavy metals. MT is a cytosolic protein in resting cells, but it can be translocated transiently to the cell nucleus during cell proliferation and differentiation [7].
Immunohistochemistry (IHC) identifies the expression, intracellular localization as well as tissue distribution of various proteins, while morphology of tissue can also be assessed precisely [8]. IHC detection of MT in tissue samples is a very important option to study its role in the pathogenesis of diseases. Special IHC methods such as multiple immunolabeling using serial sections of tissue blocks or double staining technique provide an opportunity to study correlations between MT expression and important cell and tissue functions [9]. Tissue microarray allows simultaneous examination of large number of tissues on a single microscope slide; therefore it is very suitable to evaluate the diagnostic, prognostic or predictive role of the MT expression [9].

Figure 1: Outline of standard immunohistochemical protocol [8,11,12].

Figure 2: Expression of MT-I/II in primary cutaneous malignant melanoma cells (arrows). The keratinocytes (asterisks) of epidermal basal layer served as internal positive tissue controls. The IHC detection was based on an immunoperoxidase reaction using VIP chromogenic substrate. The slides were counterstained with methyl-green. Original magnification is x20.

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