Comparative Analysis of Three Staining Techniques for Assessing Liver Fibrosis in Biopsy Samples: A Referral University Hospital

Background: Liver fibrosis is a progressive condition characterized by excessive deposition of extracellular matrix components, particularly collagen and elastic fibers, leading to impaired liver function. Accurate histological staging of fibrosis is essential for diagnosis, prognosis, and management. While hematoxylin and eosin (H and E), Masson’s trichrome (MT), and orcein (OR) stains are commonly used for fibrosis assessment, their comparative efficiency and correlation in routine practice remain inadequately defined. Methods: This retrospective study evaluated 76 archived liver biopsy samples from patients referred to Taleghani Hospital, Tehran, Iran, between 2021 and 2023. All samples underwent H and E, MT, and OR staining. Fibrosis staging was performed using the Ishak scoring system by experienced pathologists. Statistical analysis, including Fisher’s exact test, was used to assess the correlation and agreement between staining methods, with P values < 0.05 considered significant. Results: Among the 76 patients (34.2% male, mean age 41.2 years), the highest frequency of fibrosis detection occurred at Stage 2 for MT (35.5%) and at Stage 3 for OR (23.7%). Significant differences were observed between H and E and MT ( P < 0.001), and between MT and OR ( P = 0.006) in fibrosis staging. The greatest discrepancy between MT and OR was at Stage 2 ( P = 0.003). Notably, OR staining often showed a lower stage compared to MT (36.8% of cases), while in 23.7% of cases, OR indicated a higher stage. Gender and age group analyses revealed significant differences between MT and OR in males ( P = 0.036) and patients aged 0–32 years ( P = 0.032). Conclusions: Our findings demonstrate that MT and OR staining methods provide complementary information in the assessment of liver fibrosis. The significant discrepancies observed, especially at intermediate stages, suggest that incorporating OR staining alongside MT enhances diagnostic accuracy. Routine use of both stains is recommended for comprehensive fibrosis evaluation in liver biopsies.