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Nestin is a protein expressed in neural crest cells, CNS neural stems cells, as well as a few non-neural cell types in the embryo, including cells within the pancreatic islets of Langerhans and the limb bud. Nestin is an intermediate filament-associated protein, and contributes to the cytoskeleton.
Image: Nestin staining of human week-11 fetal neural progenitor cells. Cells were stained using goat anti-mouse Alexa Fluor 488 secondary antibody (green) (Molecular Probe, A-11001) and counterstained with DAPI (blue).
Image: Nestin staining of human neural progenitor cells grafted into the rat brain, 24 weeks post-op. One is of processes running along the host corpus callosum, one of cells in the hippocampus, weakly counterstained with human nuclear antigen (brown). Nestin concentration was 1:1000, overnight at room temp, visualised using the DAKO vector SG kit. Courtesy of Dr. Rike Zietlow, Cardiff University
Image: Nestin staining of human glioblastoma cells.
Images: Immunostaining of hNP1 Progenitors before and after differentiation. The culture was highly homogenous with neural rosettes. (A) Neural rosettes (white arrows, bright field) from WA09 cells. A similar result also obtained with BG02 cell line. (B) Shown here, a neural rosette stained with Nestin (NES) (green) antibody. Propagated Neural progenitors showed expression of marker genes, NES (C) and Musashi 1 (D) but not SOX2 (E). Further differentiation produced neurons (Tuj1) (F), astrocytes (GFAP) (G) and oligodendrocytes (myelin basic protein)(H) (lower panel). DAPI (blue) was used for staining the nuclei (scale bar for (A) through (E) is 100 mm and for the remaining figures 10 mm). Differentiation (2007) DOI: 10.1111/j.1432-0436.2007.00256.x...Dilutions: NES (1:100, Neuromics, Edina, MN), MSI1 (1:100, Neuromics), Tuj1 (1:500, Neuromics)...
Images: Accurate detection of intracellular antigens with optimized fixation-permeabilization conditions preserving surface antigens. Flow cytometric detection of TUJ1, MAP2 and nestin antigens in BJ fibroblasts and the neural SH-SY5Y cell line (A). TUJ1 and nestin are present in both cell lines, while the mature neuronal marker MAP2 was only detected in SH-SY5Y cells (arrows). Note stable fluorescent levels of the negative population, indicating low background staining using this protocol. Representative experiment of three independent repeats shown. (B) Corresponding validation by immunofluorescence analysis. (C) Quantitation of TUJ1, MAP2 and nestin intracellular antigen detection (n=3). Error bars indicate standard deviation. (D) Response of TUJ1 and MAP2 intracellular antigen expression to 6 DIV of 10 µM retinoic acid (RA) treatment of SH-SY5Y cells. Note disappearance/reduction of subsets negative for these markers (upward shift, green arrows), as well as a shift toward CD184low expression with differentiation (blue arrows). doi:10.1371/journal.pone.0068519.g003