The InSituPlex® Staining Method for Multiplexed Immunofluorescence Cell Phenotyping and Spatial Profiling of Tumor
Mael Manesse1, Katir K Patel1, Mark Bobrow1
1Ultivue, Cambridge, MA, USA.
Insights
The InSituPlex method offers rapid spatial profiling of immuno-oncology targets in FFPE tumor tissue. This multiplexed immunohistochemistry assay efficiently analyzes protein expression and cell interactions within the tumor microenvironment.
Area of Science:
- Oncology
- Immunology
- Pathology
- Biotechnology
Background:
- Multiplexed immunohistochemistry (mIHC) is crucial for understanding disease at the cellular level.
- Current mIHC methods are time-consuming, require specialized equipment, and involve complex optimization.
- These limitations hinder the widespread adoption of mIHC in routine diagnostics.
Purpose of the Study:
- To demonstrate the utility of the InSituPlex method for spatial profiling of immuno-oncology targets.
- To evaluate the UltiMapper™ I/O PD-L1 multiplex assay for analyzing FFPE tumor tissues.
- To assess the speed and efficiency of the InSituPlex workflow.
Main Methods:
- Utilized the InSituPlex method with the UltiMapper™ I/O PD-L1 multiplex assay.
- Analyzed Formalin-Fixed Paraffin-Embedded (FFPE) tumor tissues.
- The assay included five protein markers: CD8, CD68, PD-L1, pan CK, and SOX10.
Main Results:
- The InSituPlex method enabled detection of high and low marker expression, coexpression, and colocalization of proteins in single cells.
- Staining and image acquisition were completed in 5.5 hours.
- Demonstrated multiplexed characterization of protein expression in whole tissue sections.
Conclusions:
- The InSituPlex method provides a fast and robust workflow for deep phenotyping of tumors and their microenvironments.
- The UltiMapper™ I/O PD-L1 multiplex assay is compatible with existing instrumentation.
- This approach facilitates routine use of mIHC for immuno-oncology research and diagnostics.
Abstract:
Multiplexed immunohistochemistry (mIHC) enables the detection, quantification, and localization of many markers within cell or tissue samples, leading to a better understanding of the architecture of a disease at the cellular level. Current mIHC techniques involve long staining and assay times, require dedicated and/or captive instrumentation, and entail tedious assay optimization, hindering their establishment as routine methods. Here, we demonstrate the use of the InSituPlex® method for spatial profiling of immuno-oncology targets in FFPE tumor tissue with the UltiMapper™ I/O PD-L1 multiplex assay. The panel consists of five protein markers to profile immune infiltration and PD-L1 expression and includes CD8, CD68, PD-L1, pan CK, and SOX10 markers. The assay shows benefits of high and low expression of markers, coexpression and colocalization of proteins in single cells, and completion of staining and image acquisition in 5.5 h. Through the combination of multiplexed characterization of protein expression in whole tissue sections, fast staining workflow, and compatibility with existing instrumentation, the InSituPlex method provides a robust modality for deep phenotyping of the tumor and its microenvironment.
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