The Society for Immunotherapy of Cancer statement on best practices for multiplex immunohistochemistry (IHC) and immunofluorescence (IF) staining and validation

Abstract

ObjectivesThe interaction between the immune system and tumor cells is an important feature for the prognosis and treatment of cancer. Multiplex immunohistochemistry (mIHC) and multiplex immunofluorescence (mIF) analyses are emerging technologies that can be used to help quantify immune cell subsets, their functional state, and their spatial arrangement within the tumor microenvironment.MethodsThe Society for Immunotherapy of Cancer (SITC) convened a task force of pathologists and laboratory leaders from academic centers as well as experts from pharmaceutical and diagnostic companies to develop best practice guidelines for the optimization and validation of mIHC/mIF assays across platforms.ResultsRepresentative outputs and the advantages and disadvantages of mIHC/mIF approaches, such as multiplexed chromogenic IHC, multiplexed immunohistochemical consecutive staining on single slide, mIF (including multispectral approaches), tissue-based mass spectrometry, and digital spatial profiling are discussed.ConclusionsmIHC/mIF technologies are becoming standard tools for biomarker studies and are likely to enter routine clinical practice in the near future. Careful assay optimization and validation will help ensure outputs are robust and comparable across laboratories as well as potentially across mIHC/mIF platforms. Quantitative image analysis of mIHC/mIF output and data management considerations will be addressed in a complementary manuscript from this task force.

NK cell infiltration is associated with improved overall survival in solid cancers: A systematic review and meta-analysis

“…An overabundance of inhibitory ligands or infiltration of cells with immunosuppressive activity, including the ILC1 subset which has several overlapping features with conventional NK cells, would be expected to be associated with poor prognosis 91 . With novel multiplexing technologies and improved understanding of the distinction between NK cells and ILC subsets, it is now possible to distinguish conventional NK subsets from other populations that share markers with NK cells, including ILC1s 146 , 147 . Understanding the roles and distribution of ligands for NK cells in solid tumors will be important toward refining and precisely delivering NK-based immunotherapies.…”

Developing an enhanced 7-color multiplex IHC protocol to dissect immune infiltration in human cancers

“…Currently, 8 biomarkers can be stained together using the TSA-Opal system and the advantages and disadvantages of this method were recently reviewed [ 34 ]. There is concern regarding adding >8 markers to the panel as overheating the tissue may to excessive epitope retrieval and eventually loss of signal with consecutive rounds of staining.…”

Influence of tumor-infiltrating immune cells on local control rate, distant metastasis, and survival in patients with soft tissue sarcoma

“…First, the combination of immunohistochemical markers used to identify the immune cell phenotypes may not identify all relevant cells. However, since standardized, uniformly available assays for immunophenotyping are still pending, 43,44 this study focused on established markers of B-cells, T-cells and macrophages with a multiplex IHC staining platform. Second, we used CD68 as a general marker for macrophages staining for both M1-and M2-like subtypes, 31,45,46 rather than further differentiating between these subtypes by applying additional markers.…”