A modified Spalteholz technique with preservation of the histology

The Spalteholz technique for producing clear macroscopic specimens was intr oduced 100 years ago, and is still in use today because it immediately prov ides transparent three-dimensional specimens. The classical clearing proced ure with peroxide severely damages formalin-fixed tissue and inhibits furth er investigation on a histological level. We, therefore, wanted to make a t ransparent specimen by a modified Spalteholz method and chose the human lar ynx as an example of a large specimen. Its blood vessels were injected with gelatin, the specimen treated with acetone and peracetic acid and subjecte d to freeze substitution. After complete dehydration, benzyl benzoate and d imethylphthalate were used for impregnation. The cleared specimen satisfact orily revealed the laryngeal blood vessels and such structures as muscles, cartilage and the thyroid gland. The histology of the laryngeal glands, thy roid tissue, and infrahyoid muscle fibres was preserved. Tissue shrinkage b ecame apparent with subsequent steps of the clearing procedure. We conclude that our modifications maintain the transparency of the specimen and allow histological investigation. The herein described technique thus constitute s an improvement of the classical Spalteholz technique.