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Updated: Aug 12, 2026
Automated Multiplex Immunofluorescence Panel for Immuno-oncology Studies on Formalin-fixed Carcinoma Tissue Specimens
Published on: January 21, 2019
Utilisation of cytological samples for multiplex immunofluorescence assay
Laura Garcia Tobar1, María Villalba-Esparza1,2, Marta Abengozar-Muela1,3
1Department of Pathology, Clínica Universidad de Navarra, Pamplona, Spain.
Insights
Multiplex immunofluorescence (mIF) assays can effectively analyze immune cells in non-small cell lung cancer (NSCLC) cytological samples. This minimally invasive method aids in understanding the tumor immune microenvironment for developing novel immunotherapies.
Area of Science:
- Oncology
- Immunology
- Pathology
Background:
- Understanding the tumor immune microenvironment is crucial for developing effective non-small cell lung cancer (NSCLC) immunotherapies.
- Characterizing tumor-infiltrating immune cells and their interactions is essential for therapeutic advancements.
Purpose of the Study:
- To evaluate the utility of multiplex immunofluorescence (mIF) assays for characterizing immune cells within NSCLC cytological samples.
- To assess the feasibility of using mIF for exploring the immune landscape of NSCLC.
Main Methods:
- Six NSCLC cytological samples were analyzed using a custom mIF assay.
- The assay detected key immune markers (CD3, CD8, CD20, CD11b, CD68) and NSCLC cells (pan-cytokeratin).
- Image acquisition was performed on a Vectra-Polaris Automated Quantitative Pathology Imaging System.
Main Results:
- The mIF assay reliably detected and quantified myeloid cells (CD11b, CD68), T cells (CD3+, CD8+), and B lymphocytes (CD20+) in NSCLC cytological samples.
- Whole-tissue analysis correlated with H&E stains, improving understanding of tissue morphology and tumor-stroma interactions.
- Consistent and specific staining patterns were observed for all immune markers.
Conclusions:
- Multiplex immunofluorescence assays are feasible for analyzing immune cells in minimally invasive NSCLC cytological samples.
- This approach enables comprehensive exploration of the NSCLC immune microenvironment.
- mIF assays hold promise for guiding the development of targeted immunotherapies.
Objective:
Understanding the immune environment of non-small cell lung cancer (NSCLC) is important for designing effective anticancer immunotherapies. We describe the use of multiplex immunofluorescence (mIF) assays to enable characterisation of the tumour-infiltrating immune cells and their interactions, both across and within immune subtypes.
Methods:
Six cytological samples of NSCLC taken by transoesophageal ultrasound-guided fine needle aspiration were tested with an mIF assay designed to detect the expression of key immune cell markers such as CD3, CD8, CD20, CD11b, and CD68. Pan-cytokeratin was used to detect the NSCLC cells. Fluorescence images were acquired on a Vectra-Polaris Automated Quantitative Pathology Imaging System (Akoya Biosciences).
Results:
MIF assay was able to reliably detect and quantify the myeloid cell markers CD11b, CD68, CD3+ and CD8+ T cells, and CD20+ B lymphocytes on cytological samples of NSCLC. Whole-tissue analysis and its correlation with the corresponding H&E stains allowed a better understanding of the tissue morphology and the relationship between tumour and stroma compartments. Additionally, a uniform, specific, and correct staining pattern was seen for every immune marker.
Conclusion:
The implementation of mIF assay on cytological samples taken with minimally invasive methods seems feasible and can be used to explore the immune environment of NSCLC.
