Image Analysis of Eosinophil Peroxidase Immunohistochemistry for Diagnosis of Eosinophilic Esophagitis

Abstract

Background Diagnosis of eosinophilic esophagitis (EoE) requires manual quantification of tissue eosinophils. Eosinophil peroxidase (EPX) is an eosinophil-specific, cytoplasmic granule protein released during degranulation. Aims The objective of this study was to evaluate image analysis of EPX immunohistochemistry as an automated method for histologic diagnosis of EoE. Methods We performed a secondary analysis of prospectively collected esophageal biopsies obtained from adult subjects with EoE and controls. Tissue sections were stained with hematoxylin and eosin (H&E) and evaluated for peak eosinophils per high power field (eos/hpf). The same slides were de-stained and re-stained to detect EPX for direct comparison. Slides were digitized, and EPX staining area/mm² was quantified using image analysis. Paired samples were compared for changes in EPX staining in treatment responders and non-responders. Results Thirty-eight EoE cases and 49 controls were analyzed. Among EoE subjects, matched post-treatment biopsies were available for 21 responders and 10 non-responders. Baseline EPX/mm² was significantly increased in EoE subjects and decreased in treatment responders. EPX quantification correlated strongly with eos/hpf (r = 0.84, p < 0.0001) and identified EoE subjects with high diagnostic accuracy (AUC 0.95, p < 0.0001). The optimal diagnostic EPX-positive pixel/area threshold was 17,379 EPX/mm². Several controls (5/49) with < 15 eos/hpf on H&E staining exceeded this cutoff. Conclusions EPX/mm² correlates strongly with eos/hpf, accurately identifies subjects with EoE, and decreases in treatment responders. Automated quantification of intact eosinophils and their degranulation products may enhance pathologic assessment. Future studies are needed to correlate EPX/mm² with symptoms, endoscopic findings, and esophageal distensibility.