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Updated: Sep 30, 2025
Diagnosis of Hirschsprung's Disease by Immunostaining Rectal Suction Biopsies for Calretinin, S100 Protein and Protein Gene Product 9.5
Published on: April 26, 2019
Calretinin immunohistochemical staining in Hirschsprung's disease: An institutional experience
Ebru Zemheri1, Pinar Engin Zerk2, Cigdem Ulukaya Durakbasa3
1Department of Pathology, University of Health Sciences, Umraniye Training and Research Hospital, Istanbul, Turkey.
Insights
Calretinin staining is a reliable diagnostic tool for Hirschsprung
Area of Science:
- Gastroenterology
- Pathology
- Pediatric Surgery
Background:
- Hirschsprung's disease (HD) diagnosis relies on identifying aganglionic segments of the colon.
- Accurate histological assessment of nerve fibers and ganglion cells is crucial for diagnosis.
Purpose of the Study:
- To evaluate the efficacy of calretinin staining in diagnosing Hirschsprung's disease (HD).
- To assess the correlation between calretinin staining patterns and established diagnostic criteria for HD.
Main Methods:
- Retrospective analysis of 96 tissue samples from 56 patients over 3 years.
- Calretinin immunostaining evaluated nerve fibers and ganglion cells in various layers of intestinal tissue.
- Pathological diagnosis based on ganglion cell presence (G0/G1) and nerve hypertrophy (N0/N1).
Main Results:
- Calretinin staining was negative in aganglionic samples (G0N0) and positive in samples with ganglion cells (G1N0).
- Strong positive calretinin staining (C1) showed a significant association with G1N0 (OR: 131.401, p<0.001).
- Weak positive staining (C2) was also significantly associated with G1N0 compared to G0N0 (OR: 37.575, p=0.006).
Conclusions:
- Calretinin staining is consistently positive in the presence of ganglion cells, irrespective of nerve hypertrophy or sample characteristics.
- Calretinin staining is negative in all aganglionic samples, confirming its utility.
- Calretinin immunostaining serves as a dependable ancillary method for diagnosing Hirschsprung's disease.
Objective:
This study aims to evaluate the results obtained by calretinin staining on tissue samples for diagnosing Hirschsprung's disease (HD) in a single institution, by single expert.
Methods:
A retrospective evaluation was done for calretinin immunostaining in HD patients for a period of 3 years. Calretinin staining was evaluated in nerve fibers. Calretinin immunohistochemistry was considered positive if any staining was seen in nerve fibers and/or ganglion cells in the lamina propria, muscularis mucosa or submucosa. According to staining intensity, staining was classified as strong, weak or negative. The pathological diagnosis was based on presence or absence of ganglion cells (G0/G1) and nerve hypertrophy (N0/N1). Samples were classified according to the depth (presence of submucosa or intermuscular area), the type (biopsy or resection specimen) and staining intensity of calretinin (strong, weak, or negative staining).
Results:
A total of 96 tissue samples from 56 patients were studied. Tissues were from colon (43.8%), rectum (43.8%), stoma (6.2%), ileum (3.1%) and appendix (3.1%). The pathological diagnosis was G0N0 in 14.6%, G1N0 in 54.2%, G0N1 in 25% and G1N1 in 6.2% of cases. Our materials consisted of 92 tissue biopsies and four resection specimens. Intermuscular layer was present in 87.5% of materials and 12.5% of biopsies contained submucosa. Calretinin staining was negative (C0) in 37.5% of cases, strong positive (C1) in 47.9%, and weak positive (C2) in 14.6%. When the C0 category was taken as the reference, the status of calretinin staining as C2 (weak positive) in cases with pathological diagnosis of G1N0 was found to be 37.575 times that of cases with G0N0 (OR [95% CI]: 37.575 [2.928, 482.176], p=0.006) and the status of calretinin staining as C1 (strong positive) in cases with pathologic diagnosis of G1N0 was found to be 131.401 times that of G0N0 (OR [95% CI]: 131.401 [9.263, 1864.082), p<0.001).
Conclusion:
Calretinin staining is positive whenever ganglion cells are present independent from presence of nerve hypertrophy, the depth and the site of the biopsy or staining intensity. It is negative in all aganglionic samples. Calretinin staining is a reliable ancillary test in HD diagnosis.
