Immunohistological Markers

Double labeling with BrdU and NeuN: This image represents a section of a rat hippocampal tissue where cells are stained for neuronal (NeuN, red) and proliferation markers (BrdU, green; Ki67 blue).

The process of neurogenesis occurs in several different stages, beginning with proliferation of early progenitor cells, and ending with synaptic integration, where the newly generated cell becomes a functionally integrated neuron.  At different time-frames, various markers are expressed that correspond with the separate steps of the process. While different markers label for different things, certain markers tend to overlap one-another, so one must use precaution when drawing any definitive conclusions.  In order to avoid some of these confounds, a double labeling technique is used so as not to confuse specific cell morphologies. For example, the majority of research designs regarding neurogensis use the double labeling technique with BrdU and NeuN.  BrdU labels for newly generated cells, and NeuN is a specific label for adult neurons. If a cell expresses both of these labels, the results are more definitive and the use of specific markers allows for the advancement of scientific research to investigate the time course and fate of neurons during adult neurogensis. Below is a list of specific markers and what they label for.

Individual Markers

GFAP: astrocytes and progenitor cells

Nestin: neuronal stem cells/known to re-express itself in glial cells

Pax6: newly generated cells during the differentiation phase/also known to label hilar mature neurons/certain astrocytes

NeuroD: adult neurons

Positive stain for GFAP

Positive stain for GFAP

PSA-NCAM: progenitor cells related to neural stem cells

TUC-4: late mitotic neuronal progenitor cells and early post mitotic neurons

TuJ-1: immature neurons

NeuN: post-mitotic neurons

Calbindin: cerebellar purkinje cells and granule cells

Doublecortin: adult neurons

Fig. 2 in von Bohlen Und Halbach O. (2007). Immunohistological markers for staging neurogenesis in adult hippocampus. Cell and Tissue Research. 329(3), 409-20.

Positive staining for nestin (green), a neural stem cell marker, Photo courtesy: John Butler, Dr. Jeanne Lawrence, Dept of Cell Biology, UMASS Medical School

Faiz M, Acarin L, Castellano B, & Gonzalez B. (2005). Proliferation dynamics of germinative zone cells in the intact and excitotoxically lesioned postnatal rat brain. BMC Neuroscience. 6, 26.

Double leabeling with BrdU (blue) and doublecortin (red)

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