89 Immunohistochemistry panel design with directly-conjugated antibodies for spatial characterization of the tumor immune microenvironment
Year: 2026
Type: conference-abstract
Abstract: Background Spatial biology approaches are increasingly used to characterize cellular organization and immune contexture within the tumor microenvironment. Directly-conjugated primary antibodies for multiplex immunohistochemistry (IHC) offer a streamlined alternative to amplification-based workflows, reducing assay complexity while enabling simultaneous detection of multiple biomarkers. However, panel design and optimization remain critical for generating reproducible, high quality spatial data.Methods We optimized a multiplex spatial biology workflow utilizing directly-conjugated primary antibodies for analysis of healthy colon and colon adenocarcinoma tissues. The workflow incorporated in-house antibody conjugation, strategies to minimize tissue autofluorescence, and spectral unmixing optimization to improve signal discrimination. A colorectal cancer immuno-oncology panel was designed to evaluate immune, epithelial, and proliferation-associated markers, including CD3, CD8, CD68, pan-cytokeratin (Pan-CK), p53, Ki-67, and β-catenin.Results The optimized workflow enabled robust multiplex detection and spatial resolution of immune and tumor-associated biomarkers within formalin-fixed paraffin-embedded tissue sections. Comparative analysis of healthy colon and colon adenocarcinoma samples identified differences in marker distribution and tissue architecture, including reduced abundance of immune cell markers and uniform p53 expression within tumor regions. Spectral unmixing and autofluorescence reduction improved marker separation and overall image quality, facilitating accurate spatial characterization of the tumor microenvironment.Conclusions Directly-conjugated primary antibody panels provide a scalable and reproducible approach for spatial profiling of colorectal tissues. The described framework supports efficient panel development and optimization for investigation of immune and structural features within the tumor microenvironment and may facilitate biomarker discovery and translational immuno-oncology research.For Research Use Only.
Authors: Jen Serafin, Mae Voeun, Kristina Hamilton
Institution: Thermo Fisher Scientific (United States)
Language: English
Subfield: Oncology
Field: Medicine
Domain: Health Sciences
Open Access status: gold
Estimated APC paid: $2,490
OpenAlex ID: W7225184490