Enzyme and immunohistochemistry on undecalcified bone and bone marrow biopsies after embedding in plastic

A simplified method of low temperature methyl and butyl methacrylate embedding (up -20 degrees to -15 degrees C) is demonstrated using a proper redox system of benzoyl peroxide and aromatic amine. This method combines the morphological superiority of plastic-embedded bone tissue and bone marrow sections with the advantages of specific enzyme histochemical and immunochemical markers. The method permits good preservation of morphological details, the survival of antigenic determinants and the retention of enzyme activities. The specimens were fixed in 1.6% formaldehyde and 5% sucrose in 0.02 M phosphate buffer at pH 7.4, washed in 0.02 M phosphate buffer and 5% sucrose, dehydrated with acetone and impregnated with monomers of embedding medium. All these steps were carried out at +4 degrees C. The method presented is especially suitable for enzyme histological and immunohistological diagnosis of primary and secondary bone tumours, soft tissue tumours, as well as myelo- and lymphoproliferative disorders of bone marrow biopsies. Examples are demonstrated with mono- and polyclonal antibodies and reaction products of hydrolytic enzymes.