Multiplex immunohistochemistry (IHC) profiling of murine tumour-infiltrating T cells

The tumour microenvironment (TME) represents a heterogenous and dynamic niche, comprising cancer cells, extracellular matrix, stromal cells, vasculature and immune cells. The composition of the immune infiltrate varies between patient and tumour type, with the overall number, spatial localisation and functional status constituting the immune contexture. The presence of certain immune cells, including specific subsets of CD8 and CD4 T cells, natural killer cells, B cells and dendritic cells has been linked with favourable prognosis and response to treatment, including immunotherapy. In contrast, subsets of immune cells including macrophages, myeloid cells and neutrophils are frequently associated with poor prognosis, treatment resistance and tumour growth. Therefore, assessment of the immune contexture can be used to evaluate prognostic and predictive immune biomarkers, with CD8 cytolytic and memory T cells emerging as key immune effectors associated with clinical benefit across multiple solid malignancies. Here, we illustrate a straightforward automated immunohistochemistry (IHC) protocol for the detection and characterization of different subtypes of T-cells within the microenvironment of murine tumours. We also briefly illustrate the downstream process of visualization of these specific lymphocytic populations, ultimately providing both qualitative and quantitative analysis. The general protocol can readily be adapted to study a wide range of murine tumours and normal tissue (including secondary lymphoid organs), as well as profiling of other immune cell populations.