Double Labeling of Proliferating Neurons with Anti-BrdU and Anti-NeuN: An Improved Immunohistochemical Technique Utilizing Microwave Irradiation

The authors describe a double labeling immunohistochemical technique to facilitate the study of neuronal proliferation and migration. Proliferating fetal neuroblasts were labeled with 5-bromo-2′-deoxyuridine (BrdU), and the labeled mature neurons were identified with a monoclonal anti-BrdU antibody. To differentiate neurons from glia, the tissue was also treated with monoclonal anti-NeuN, an antibody that detects a nuclear antigen unique to neurons. Novel aspects of this procedure are the utilization of methacarn fixation of tissues and the denaturation of BrdUlabeled DNA with microwave irradiation rather than with either acid or alkali denaturation treatments. This method produces strong immunoreactivity for both BrdU labeled neurons and neurons reacting with anti-NeuN, thereby facilitating the quantitative study of the development of cortical cytoarchitecture. (The J Histotechnol 21:201–204, 1998)