Comparison of aldehyde and ethanol fixatives in urogenital tissue histology

Formaldehyde remains widely used as a fixative despite its documented risks. Ethanol-based fixatives have emerged as safer alternatives, but their performance in urogenital tissues has not been systematically assessed.

To compare the histomorphological preservation of rat kidneys, prostate, penis, and testis fixed with ethanol versus formaldehyde-based solutions.

Six adult male rats were euthanized, and urogenital organs were collected and sectioned. Testes were initially fixed in Bouin's solution for 24h. All tissues were subsequently immersed in buffered formaldehyde (BF), buffered paraformaldehyde (BP), 70% unbuffered ethanol (UE), or 70% buffered ethanol (BE) for 96h (or 72h for testes following Bouin). Samples were processed for paraffin embedding and stained with H&E. Four blinded examiners scored tissue preservation, staining quality, and artifacts using a 1-5 scale. Data were analyzed using Kruskal-Wallis with Dunn's post-test (p<0.05).

Ethanol-based fixatives (UE and BE) consistently outperformed aldehyde-based fixatives (BF and BP). In kidneys, UE and BE yielded fewer artifacts and superior preservation of glomerular and tubular structures. In the prostate, these fixatives provided better epithelial and stromal quality and fewer artifacts. All fixatives adequately preserved penile cavernous tissue, but BE showed superior urothelial integrity. Following Bouin's pre-fixation, UE and BE improved staining and preservation of the germinal epithelium and Sertoli cells compared with aldehyde fixatives.

Ethanol-based fixatives provide superior or comparable histomorphological preservation of urogenital tissues while avoiding the health and environmental risks associated with formaldehyde.