Diagnostic assessment of Hirschsprung disease using fluorescence confocal microscopy: A feasibility study

INTRODUCTION: Hirschsprung disease (HD) is a congenital disorder marked by aganglionosis of intestinal nerve plexuses, leading to bowel obstruction. Determining the extent of the aganglionic segment is essential for successful surgical correction. Consequently, the extent of aganglionosis must be examined histologically. Fluorescence confocal microscopy (FCM) may serve as an alternative diagnostic modality. Using a laser to scan the unfixed specimen, it enables rapid intraoperative imaging without causing tissue alteration. This study assessed the feasibility and diagnostic accuracy of FCM in evaluating Hirschsprung disease in pediatric patients. MATERIALS AND METHODS: Patients undergoing rectal biopsy, ostomy closure, or transanal endorectal pull-through (TERPT) at our center from January to August 2024 were included. Tissue samples were imaged by FCM followed by frozen section and formalin-fixed paraffin-embedded (FFPE) histology. Image quality, assessability of submucosal and myenteric plexuses, and diagnostic agreement with FFPE sections were investigated. RESULTS: A total of 34 samples from 8 patients were analyzed by FCM and FFPE, the 24 TERPT-derived samples were additionally analyzed by frozen section. Diagnostic agreement with FFPE was 78.5 % for FCM and 86.9 % for frozen sections (p = 0.49). Among fully assessable samples, FCM achieved 80.9 % agreement with FFPE compared to 86.4 % for frozen sections. CONCLUSION: FCM is an easy-to-implement method for identifying normal and affected bowel in HD. It is used on unfixed specimens, preserving the tissue quality. Its diagnostic accuracy is comparable to that of frozen sections in assessable samples, and it is advantageous in cases where local access to expert pathologists is limited.