Faculty, Staff and Student Publications
Publication Date
1-8-2025
Journal
The Journal for ImmunoTherapy of Cancer
Abstract
Objectives: Multiplex immunohistochemistry and immunofluorescence (mIHC/IF) are emerging technologies that can be used to help define complex immunophenotypes in tissue, quantify immune cell subsets, and assess the spatial arrangement of marker expression. mIHC/IF assays require concerted efforts to optimize and validate the multiplex staining protocols prior to their application on slides. The best practice guidelines for staining and validation of mIHC/IF assays across platforms were previously published by this task force. The current effort represents a complementary manuscript for mIHC/IF analysis focused on the associated image analysis and data management.
Methods: The Society for Immunotherapy of Cancer 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 quantitative image analysis of mIHC/IF output and data management considerations.
Results: Best-practice approaches for image acquisition, color deconvolution and spectral unmixing, tissue and cell segmentation, phenotyping, and algorithm verification are reviewed. Additional quality control (QC) measures such as batch-to-batch correction and QC for assembled images are also discussed. Recommendations for sharing raw outputs, processed results, key analysis programs and source code, and representative photomicrographs from mIHC/IF assays are included. Lastly, multi-institutional harmonization efforts are described.
Conclusions: mIHC/IF technologies are maturing and are routinely included in research studies and moving towards clinical use. Guidelines for how to perform and standardize image analysis on mIHC/IF-stained slides will likely contribute to more comparable results across laboratories and pave the way for clinical implementation. A checklist encompassing these two-part guidelines for the generation of robust data from quantitative mIHC/IF assays will be provided in a third publication from this task force. While the current effort is mainly focused on best practices for characterizing the tumor microenvironment, these principles are broadly applicable to any mIHC/IF assay and associated image analysis.
Keywords
Humans, Fluorescent Antibody Technique, Image Processing, Computer-Assisted, Immunohistochemistry, Immunotherapy, Neoplasms, Education, Immunotherapy, Pathology, Tumor microenvironment - TME
DOI
10.1136/jitc-2024-008875
PMID
39779210
PMCID
PMC11749220
PubMedCentral® Posted Date
1-8-2025
PubMedCentral® Full Text Version
Post-print
Published Open-Access
yes
Included in
Bioinformatics Commons, Biomedical Informatics Commons, Genetic Phenomena Commons, Medical Genetics Commons, Oncology Commons