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Impact of Region-of-Interest Size on Immune Profiling Using Multiplex Immunofluorescence Tyramide Signal Amplification for Paraffin-Embedded Tumor Tissues.


ABSTRACT:

Introduction

Representative regions of interest (ROIs) analysis from the whole slide images (WSI) are currently being used to study immune markers by multiplex immunofluorescence (mIF) and single immunohistochemistry (IHC). However, the amount of area needed to be analyzed to be representative of the entire tumor in a WSI has not been defined.

Methods

We labeled tumor-associated immune cells by mIF and single IHC in separate cohorts of non-small cell lung cancer (NSCLC) samples and we analyzed them as whole tumor area as well as using different number of ROIs to know how much area will be need to represent the entire tumor area.

Results

For mIF using the InForm software and ROI of 0.33 mm2 each, we observed that the cell density data from five randomly selected ROIs is enough to achieve, in 90% of our samples, more than 0.9 of Spearman correlation coefficient and for single IHC using ScanScope tool box from Aperio and ROIs of 1 mm2 each, we found that the correlation value of more than 0.9 was achieved using 5 ROIs in a similar cohort. Additionally, we also observed that each cell phenotype in mIF influence differently the correlation between the areas analyzed by the ROIs and the WSI. Tumor tissue with high intratumor epithelial and immune cells phenotype, quality, and spatial distribution heterogeneity need more area analyzed to represent better the whole tumor area.

Conclusion

We found that at minimum 1.65 mm2 area is enough to represent the entire tumor areas in most of our NSCLC samples using mIF.

SUBMITTER: Sun B 

PROVIDER: S-EPMC9684353 | biostudies-literature | 2023

REPOSITORIES: biostudies-literature

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Publications

Impact of Region-of-Interest Size on Immune Profiling Using Multiplex Immunofluorescence Tyramide Signal Amplification for Paraffin-Embedded Tumor Tissues.

Sun Baohua B   Laberiano-Fernández Caddie C   Salazar-Alejo Ruth R   Zhang Jiexin J   Solorzano Rendon Jose Luis JL   Lee Jack J   Solis Soto Luisa Maren LM   Wistuba Ignacio Ivan II   Parra Edwin Roger ER  

Pathobiology : journal of immunopathology, molecular and cellular biology 20220524 1


<h4>Introduction</h4>Representative regions of interest (ROIs) analysis from the whole slide images (WSI) are currently being used to study immune markers by multiplex immunofluorescence (mIF) and single immunohistochemistry (IHC). However, the amount of area needed to be analyzed to be representative of the entire tumor in a WSI has not been defined.<h4>Methods</h4>We labeled tumor-associated immune cells by mIF and single IHC in separate cohorts of non-small cell lung cancer (NSCLC) samples an  ...[more]

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