Publication
Recording and analysing DNA from osteocytes in resin-embedded bone samples
Publisher:
Springer Science and Business Media LLC
Date:
19-11-2022
DOI:
10.1007/S12024-022-00559-1
Abstract: We report on a process to record the presence and the location of osteocyte nuclei using two nucleic staining dyes, Diamond™ Nucleic Acids Dye (DD) and DAPI (4',6-diamidino-2-phenylindole). Knowledge of the presence and number of osteocytes is key to any success in subsequent DNA profiling. Osteocytes are most numerous cells and thus the main source of DNA in bone s les, which can be preserved for histological analyses. Archived s les are either fixed in formalin or preserved in ethanol prior to embedding in resin. These resin-embedded s les are potentially used as ante mortem reference s les. Cases of a missing person investigation are one ex le where this type of preserved reference material may be of value. When resin is required for s le preservation it represents a problem for subsequent DNA profiling, if needed as a reference s le in human identification. It is essential therefore to remove the resin prior to DNA analyses as resin is a known inhibitor of DNA profiling. Current methods of resin removal are lengthy and require toxic chemicals. This report describes a simplified process to remove resin and visualise the location of nucleated osteocytes. Eight sections of bone s les at 5-µm thickness were stained with DD and DAPI. A further three s les were processed using a formalin-fixed method and three additional s les treated following an ethanol-preserved method (11 s les for both the formalin-fixed and 11 for the ethanol-preserved with eight in common). The location and number of nuclei could be recorded clearly due to the fluorescence created by the dye binding to DNA. The number of stained nuclei correlated with the mass of DNA isolated from the sections (r = 0.873, p = 1.21 × 10