Immune system cell subsets of lymph nodes and spleen from chamois (Rupicapra rupicapra subspecies rupicapra) living in the Italian Alps, have been characterised by immunohistochemical techniques. A total of seven primary antibodies (against human CD3, CD79αcy, CD68, or ovine CD4, CD8, CD21, and γδ TCR epitopes) were tested on tissues fixed either in formalin or in zinc salts (ZS) and a good cross-reactivity with chamois immune cells epitopes has been detected. ZS fixation was used to allow a wider identification of immune cells, without requirement of antigen retrieval treatments. A higher percentage of cells were labelled using this fixative furthermore CD4 and CD21 cells showed labelling only in ZS fixed tissues. However, antibodies raised against human CD3, CD79 and CD68 antigens, successfully reacted with chamois immune cells, both in ZS fixed tissues and in formalin, but in the latter antigen retrieval had to be used. CD3-positive cells were prevalent in the interfollicular/paracortical area of the lymph node and periarterial sheath of the spleen, whereas CD79 positive cells were mainly localized in the germinal centre and mantle zone of follicles both in lymph nodes and in spleen. High percentages of CD68-positive cells were detected in medullary cords of lymph node and marginal zone and red pulp of the spleen. Distribution of CD4-positive cells mirrored the distribution of CD3-positive cells and CD21-positive cells had a similar distribution to CD79-positive cells but also included follicular dendritic cells. Results from this study may allow future investigation of the immune response and pathogenesis of chamois diseases.