Processing and Sectioning Undecalcified Murine Bone Specimens

Methods Mol Biol. 2021:2230:231-257. doi: 10.1007/978-1-0716-1028-2_14.

Abstract

Preparation of mineralized tissue specimens for bone-specific staining encompasses a critical sequence of histological techniques that provides visualization of tissue and cellular morphology. Bone specimens are fixed in 10% neutral buffered formalin (NBF), dehydrated in graded ethanol (EtOH) solutions (and optionally cleared in xylene), infiltrated and embedded in polymethyl methacrylate (methyl methacrylate or MMA), classically sliced into 4-10 micrometer (μm) sections, and stained with bone-specific histological stains such as von Kossa (with either nuclear fast red solution counterstain or MacNeal's tetrachrome counterstain), modified Goldner's trichrome, Alizarin Red S, Safranin O, and tartrate-resistant acid phosphatase (TRAP) stain. Here, we describe the tissue processing of mineralized mouse bones from dissection to staining for histological analysis by light microscopy.

Keywords: Bone; Dehydration; Embedding; Fixation; Methyl methacrylate; Sectioning; Staining.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Animals
  • Calcification, Physiologic / physiology*
  • Coloring Agents / pharmacology*
  • Histological Techniques
  • Mice
  • Skull / diagnostic imaging*
  • Staining and Labeling / methods*

Substances

  • Coloring Agents