Evaluation of Proliferation of Neural Stem Cells In Vitro and In Vivo. (1st February 2013)

• Record Type: Journal Article
• Authors: Morte, Maria Inês; Carreira, Bruno P.; Machado, Vanessa; Carmo, Anália; Nunes‐Correia, Isabel; Carvalho, Caetana M.; Araújo, Inês M.
• Editors: Bhatia, Mick; Elefanty, Andrew G.; Fisher, Susan J.; Patient, Roger; Schlaeger, Thorsten; Snyder, Evan Y.
• Is Part Of: Current protocols in stem cell biology. Volume 24(2013)
• Page Start: 2D.14.1
• Page End: 2D.14.24
• Publication Date: 2013-02-01
• Subjects: EdU -- BrdU -- proliferation -- subventricular zone -- hippocampus; Stem cells -- Laboratory manuals; Stem cells -- Physiology; Stem Cells; Stem cells; Laboratory Manual; Laboratory manuals; 571.5
• DOI: 10.1002/9780470151808.sc02d14s24
• Languages: English
• ISSNs: 1941-7322

Abstract

Abstract: This unit describes two basic protocols for the detection of the proliferation of neural stem cells (NSC). The first one addresses cell proliferation in cultures, starting with primary cell cultures isolated from the mouse subventricular zone (SVZ), in which SVZ‐derived NSC are kept in culture as neurospheres. By using this culture system, we are able to study different stages of adult neurogenesis, such as proliferation, differentiation, migration, and survival. Thus, in the first basic protocol, we describe two different techniques to evaluate cell proliferation based on EdU incorporation: (a) immunocytochemistry and (b) flow cytometry. EdU, a new thymidine analog, which is detected by a reproducible and sensitive method based on click chemistry, does not require DNA denaturation, as is the case with BrdU. Thus, co‐labeling of EdU with other specific antibodies of extracellular or intracellular targets, as well as other DNA dyes, is possible. In the second basic protocol, we describe an in vivo assay to evaluate proliferation of NSC in the dentate gyrus of hippocampus of adult mice, by both BrdU and EdU detection. With this approach, it is also possible to study different stages of adult neurogenesis, by co‐labeling thymidine analogs with other specific markers, such as doublecortin (DCX) or neuronal nuclei protein (NeuN). Curr. Protoc. Stem Cell Biol . 24:2D.14.1‐2D.14.24. © 2013 by John Wiley & Sons, Inc.