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A Double Histochemical/Immunohistochemical Staining for the Identification of Canine Mast Cells in Light Microscopy
Francesca Gobbo1, Giuseppe Sarli1, Margherita De Silva1
1Department of Veterinary Medical Sciences, University of Bologna, Ozzano dell'Emilia, 40064 Bologna, Italy.
Insights
A new double staining method combines immunohistochemistry (IHC) with toluidine blue counterstaining to simplify mast cell (MC) identification in canine skin disorders. This technique aids in distinguishing neoplastic mast cells from inflammatory cells and calculating the Ki67 index.
Area of Science:
- Veterinary Pathology
- Immunohistochemistry
- Cell Biology
Background:
- Immunohistochemistry (IHC) is crucial for diagnostics but simultaneous multi-antibody analysis is complex and costly.
- Identifying mast cells (MCs) alongside other markers in canine skin conditions presents diagnostic challenges.
Purpose of the Study:
- To develop and validate a novel double staining method combining IHC with toluidine blue counterstaining.
- To facilitate the identification of mast cells (MCs) in canine cutaneous disorders, particularly when evaluating nuclear and/or membranous markers.
Main Methods:
- Tested a combined IHC and toluidine blue staining technique on canine cutaneous mast cell tumors (MCTs), cutaneous mastocytosis, and atopic dermatitis.
- Evaluated the utility of the method for c-kit, Ki67, and cannabinoid receptor 2 markers.
Main Results:
- The double staining method effectively aids in identifying MCs and distinguishing them from inflammatory cells.
- It facilitates the calculation of the Ki67 proliferation index in canine MCTs.
- The technique is particularly useful for non-cytoplasmic markers and in cases with low MC numbers.
Conclusions:
- This IHC and toluidine blue double staining is a valuable tool for evaluating canine cutaneous disorders with MC involvement.
- It simplifies the identification of MCs, aiding in the diagnosis and characterization of MCTs and other conditions.
- The method shows potential for studying new markers in tissues containing MCs in both veterinary and human medicine.
Abstract:
Immunohistochemistry (IHC) is a widely used technique in diagnostic pathology, but the simultaneous analysis of more than one antibody at a time with different chromogens is rather complex, time-consuming, and quite expensive. In order to facilitate the identification of mast cells (MCs) during immunohistochemical analysis of membrane and/or nuclear markers, we propose a new staining method that includes the association of IHC and toluidine blue as a counterstain. To achieve this goal, we tested c-kit, Ki67, and cannabinoid receptor 2 on several cases of cutaneous canine mast cell tumors (MCTs), cutaneous mastocytosis, and atopic dermatitis. The results obtained show how this double staining technique, although limited to non-cytoplasmic markers and of little use in poorly differentiated MCTs in which MC metachromasia is hard to see, can be used during the evaluation of nuclear and/or membranous immunohistochemical markers in all canine cutaneous disorders, especially if characterized by the presence of a low number of MCs. It can help to evaluate those MCTs in which neoplastic MCs must be clearly distinguished from inflammatory cells that can infiltrate the tumor itself, in facilitating the calculation of the Ki67 index. Moreover, it can be used to study the expression of new markers in both animal and human tissues containing MCs and in MC disorders.
