Time-Dependent Autolysis in Ex Vivo–Maintained Rat Tissue in Saline: A Descriptive Study and Practical Guidance for Histopathologic Assessment

Timely tissue fixation is critical to prevent autolysis, which confounds histopathologic assessment. This study systematically mapped autolysis progression in ex vivo rat tissues under room temperature and refrigerated (4°C) storage conditions to establish an evidence-based, tissue-specific timeline for practical guidance. Thirty-six Sprague-Dawley rats were euthanized; major organs were rapidly collected and maintained in sterile saline at either room temperature or 4°C for intervals ranging from 0.5 to 16 hours before fixation in 10% neutral buffered formalin and H&E evaluation. The data provide a clear timeframe of autolytic change in rat tissues. At room temperature, most tissues exhibited minimal autolysis within the first 2 hours, and refrigeration at 4°C extended the time to autolysis. Immunohistochemistry for the endothelial marker CD31 suggested that CD31 antigenicity remained detectable in most tissues maintained at room temperature out to 16 hours with the exception of intestinal tissues demonstrating severe autolysis, indicating that advanced autolysis can compromise immunohistochemical interpretation. In conclusion, this work provides guidance for handling tissues that require temporary preservation in saline. The documented tissue-specific timelines serve as a valuable reference for pathologists.