Beyond morphology: a combined histopathological and immunohistochemical protocol for the post-mortem diagnosis of the electrical mark

The histopathological diagnosis of the electrical mark is hampered by the morphological overlap with thermal burns and abrasions and by the absence of standardized immunohistochemical criteria. This study aimed to develop and preliminarily evaluate an integrated histological and immunohistochemical protocol for the microscopic diagnosis of documented electrocution. Twenty-four full-thickness cutaneous samples from forensic cases with documented electrocution were stained with hematoxylin-eosin and processed for immunohistochemistry with Cytokeratin AE1/AE3. Six representative cases were also stained with HSP27, HSP70, and HSP90 antibodies. Histological findings were summarized as frequencies with exact 95% binomial confidence intervals (CIs). Ordinal immunohistochemical scores assigned independently by three observers were evaluated using a two-way random-effects, absolute-agreement intraclass correlation coefficient for single measurements [ICC(A,1)], pairwise weighted Cohen's kappa, and observed agreement. Epidermal and dermal coagulative necrosis, collagen homogenization, vascular dilatation/congestion, and hemorrhage/thrombosis were observed in 24/24 samples (100%; exact 95% CI 85.8-100%). Epidermal and dermal nuclear elongation and dark-stained nuclei occurred in 23/24 samples (95.8%; 95% CI 78.9-99.9%). For Cytokeratin, ICC(A,1) was 0.748 (95% CI 0.568-0.873); pairwise linearly weighted kappa values ranged from 0.437 to 0.820, and exact agreement among all three observers was 43.5%. HSP scores were identical among observers across the six selected slides (ICC and pairwise weighted kappa = 1.000; exact agreement 100%, 95% CI 54.1-100%). The combined assessment of H&E features and Cytokeratin/HSP immunoreactivity identifies a recurrent histopathological pattern that supports a provisional hierarchy of findings and can improve the standardization of microscopic assessment of electrical marks.