Beyond the first cut: evaluating CMV IHC reliability in serial gastrointestinal biopsy sections

Cytomegalovirus(CMV) is a major pathogen in immunocompromised individuals, often causing severe gastrointestinal complications. Immunohistochemistry (IHC) is widely used for CMV detection due to its high specificity, but its reliability in cases with sparse CMV-positive cells remains uncertain. This study evaluates the reproducibility of CMV IHC in gastrointestinal biopsies by assessing CMV detection across serial sections. A prospective observational study was conducted at the Pathology Institute of Sheba Medical Center between the years 2023 and 2024. Twenty-five CMV colitis cases diagnosed based on IHC were included and categorized as positive (≥ 2 CMV-positive cells) or equivocal (1 CMV-positive cell). Serial sectioning of the tissue blocks and subsequent CMV IHC staining was performed. Descriptive statistical analysis, including the Friedman test and Wilcoxon signed-rank test, was used to assess variability in CMV detection. CMV-positive cell counts varied significantly across serial sections (p = 0.047, Friedman test). While some cases remained stable, others fluctuated, transitioning from positive to equivocal or negative. Among initially positive cases, 33.3% (7/21) had at least one negative section, and 33.3% (7/21) had at least one equivocal section. Equivocal cases were highly unstable, with all showing at least one negative section and one converting to positive. Pairwise Wilcoxon tests showed no consistent significant differences. CMV IHC exhibits variability in low-positive cases, leading to diagnostic transitions. Serial sectioning enhances confidence but does not fully eliminate variability. A multimodal approach integrating IHC with molecular methods and further standardization of IHC protocols may improve CMV detection accuracy.