Simultaneous detection of TDT-mediated dUTP-biotin nick end-labeling (TUNEL)-positive cells and multiple immunohistochemical markers in single tissue sections.

We have modified the terminal deoxynucleotidyl transferase (TDT)-mediated dUTP-biotin nick end-labeling (TUNEL) method to permit the immunogold-silver intensification detection of apoptotic cells in tissue sections. Such sections can subsequently be processed for multi-labeling fluorescence microscopy, thus permitting the simultaneous detection of silver-positive apoptotic cells and three additional fluorescent signals. In this report, we combined TUNEL labeling with the fluorescent detection of bromodeoxyuridine-positive proliferating cells, growth-associated protein-43 (GAP-43) immunoreactive neurons and bisbenzimide-positive pyknotic cells in the embryonic mouse nervous system. This combination of stains allowed the simultaneous detection of proliferative, differentiated and apoptotic cells in the developing nervous system.