Detection of Bromodeoxyuridine (BrdU) in Semi-Thin Sections of Routine Electron Microscopic Specimens Digested with Trypsin

Abstract

A simplified method of immunostaining of bromodeoxyuridine (BrdU), a thymidine analogue, for routinely paraformaldehyde or glutaraldehyde and osmium tetroxide-fixed and epon embedded specimens was established using regenerating rat liver. In the case of 4% paraformaldehyde fixed tissue, only removal of the epoxy resin with 10% Na-ethoxide solution was required to detect BrdU. In glutaraldehyde and osmium tetroxide-fixed specimens, 10% Na-ethoxide treatment and additional trypsin digestion enabled the detection of BrdU. HCl hydrolysis to denature DNA in the nuclei was not essential for the BrdU immunoreaction. This procedure is one of the most simple methods to detect BrdU with much better resolution. Thus, BrdU incorporated into DNA of S-phase cell is detectable in conventionally prepared specimens for routine electron microscopy.