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Application and Validation of Semiautomatic Quantification of Immunohistochemically Stained Sections for Low Cellular
Andrea Nüesch1, Maria Paola Ferri2,3, Christine L Le Maitre1
1Division of Clinical Medicine, School of Medicine and Population Health University of Sheffield Sheffield UK.
JOR Spine
|March 10, 2025
Summary
This study introduces a QuPath script for semi-automatic cell counting in low-cellularity tissues, improving accuracy and efficiency in immunohistochemistry (IHC) analysis. The method enhances reproducibility and reduces subjectivity compared to manual cell counting.
Area of Science:
- Biomedical Imaging
- Cell Biology
- Histopathology
Background:
- Immunohistochemistry (IHC) is crucial for protein localization and quantification in tissues.
- Manual cell counting in IHC is subjective, time-consuming, and impacts reproducibility.
- Digital pathology and bioimage analysis offer solutions for more efficient and accurate quantification.
Purpose of the Study:
- To develop and validate a QuPath-based script for semi-automatic cell counting.
- To specifically address the challenges of analyzing tissues with low cellularity.
- To improve the accuracy, reproducibility, and efficiency of IHC analysis.
Main Methods:
- Developed a QuPath script and guide for semi-automatic cell counting.
- Applied the method to low-cellularity tissues like intervertebral discs and cartilage.
- Validated the script by comparing results with manual counting and assessing inter-rater reliability.
Main Results:
- No significant difference between manual and semi-automatic quantification (p=0.783, p=0.386).
- Strong correlation between methods for collagen type II (r=0.9602) and N-cadherin (r=0.9044) staining.
- High inter-rater reliability (ICC=0.853) using the semi-automatic method.
Conclusions:
- The QuPath script provides a reproducible and accurate alternative to manual IHC cell counting.
- The method is particularly effective for tissues with low cellularity and high extracellular matrix.
- This approach enhances standardization and comparability in IHC analysis across laboratories.
