Publication: Demonstration of Doublecortin Protein, a Neurogenesis Marker, at the Electron Microscopic Level
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Date
2017
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AVES PRESS LTD
Abstract
Objective: Demonstration of newly born neurons in adult brains is an important issue in terms of elucidating neurogenesis. In our study, we aimed to develop an appropriate pre-embedding microwave labeling method for the newly born neuronal marker doublecortin (DCX) protein in the hippocampal dentate gyrus (DG) region. Methods: Brains were obtained from 10-week-old C57BJ/6J male mice by perfusion fixation. Vibratome sections from brain tissues were labeled with an anti-DCX antibody using a pre-embedding microwave method. Sections stained with 3.3'-diaminobenzidine (DAB) were prepared for electron microscopic (EM) analyses using a microwave. After embedding in Epon, thin sections were obtained, observed under an electron microscope, and photographed for morphological assessments. Results: DCX labeling performed using the pre-embedding microwave method was specifically demonstrated at both light and electron microscopic levels. DCX-positive cells were localized at the subgranular zone and granular layer. Electron microscopic observations showed that DCX immunoreactivity was positive at the axons, dendrites, and somata. Conclusion: We demonstrated that the existence of DCX can be determined using the pre-embedding microwave labeling as an immunoelectron method in the DG region. Our study provides a basis for further studies on neurogenesis aiming to show DCX-immunoreactive cells using the pre-embedding DAB labeling method at the electron microscopic level.
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Keywords
Neurogenesis, doublecortin, pre-embedding microwave technique, immunoelectron microscopy, CENTRAL-NERVOUS-SYSTEM, ADULT NEUROGENESIS, HIPPOCAMPAL NEUROGENESIS, DENTATE GYRUS, NEURONS, BRAIN, MICROTUBULES, EXPRESSION, CELLS