The RNAscope® assay is a unique RNA ISH technology that identifies RNA expression at the single cell level with morphological context. Here we present the use of RNAscope® for the detection of RNA targets in the tumor microenvironment (TME) that are involved in tumor immunology and immunotherapy:
- Checkpoint markers
- Immune cell markers
- Cytokines and chemokines
- Localization of specific immune cell types (i.e., cytotoxic lymphocytes and regulatory T cells) in the TME
- Spatial relationships between different cell types in the TME
- Characterization of secreted proteins (i.e., cytokines and chemokines)
- Evaluation of immune function in TME beyond enumeration of tumor infiltrating lymphocytes
Introduction
The field of cancer immunotherapy has expanded rapidly in recent years with promising clinical results by immune checkpoint inhibitors and other therapeutic approaches such as cancer vaccines and chimeric antigen receptor (CAR) therapy. Despite the dramatic and durable responses seen in many patients, our understanding of the immune response to cancer is still limited, and we cannot reliably predict who will or will not benefit from these new interventions. To better stratify patients for immunotherapy treatments, the series of events and biomarkers involved in the cancer‑immunity cycle need to be better understand(1-3). In addition, spatially mapped expression data at the single-cell level is crucial to understanding the cellular organization and cell-to-cell interactions in the tumor and its complex microenvironment (TME).
RNAscope® is a unique RNA ISH technology that provides single-cell gene expression resolution with spatial and morphological context. The RNAscope® assay detects mRNA and long noncoding RNAs in fresh frozen, fresh fixed, and formalin-fixed paraffin-embedded (FFPE) cells and tissues by utilizing a unique double Z probe design and signal amplification strategy that allows for visualization of target RNA as a single dot, where each dot is an individual RNA molecule(4). The key benefit of the RNAscope® strategy is high sensitivity due to its amplification method and high specificity because of the double Z probe design, leading to a high signal-to-noise ratio in many tissues. In this report we examined 50 selected immune checkpoint and functional markers.
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