The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.
Use a concentration of 1 µg/ml. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
FunctionRequired for brain and eye development. Promotes the disassembly of phosphorylated vimentin intermediate filaments (IF) during mitosis and may play a role in the trafficking and distribution of IF proteins and other cellular factors to daughter cells during progenitor cell division. Required for survival, renewal and mitogen-stimulated proliferation of neural progenitor cells.
Tissue specificityCNS stem cells.
Sequence similaritiesBelongs to the intermediate filament family.
Developmental stageUpon terminal neural differentiation, nestin is down-regulated and replaced by neurofilaments.
Post-translational modificationsConstitutively phosphorylated. This increases during mitosis when the cytoplasmic intermediate filament network is reorganized.
IHC image of Nestin staining in Rat brain (6 weeks) FFPE section, performed on a BondTM system using the standard protocol F. The section was pre-treated using heat mediated antigen retrieval with sodium citrate buffer (pH6, epitope retrieval solution 1) for 20 mins. The section was then incubated with ab93157, 1µg/ml, for 15 mins at room temperature and detected using an HRP conjugated compact polymer system. DAB was used as the chromogen. The section was then counterstained with haematoxylin and mounted with DPX.
Immunohistochemistry (PFA perfusion fixed frozen sections) - Anti-Nestin antibody (ab93157)Sophie Pezet, Laboratoire de Neurobiologie, France
IHC-FoFr image of nestin staining in rat brain sections using ab93157(1:100). The sections used came from animals perfused fixed with Paraformaldehyde 4% with 15% of a solution of saturated picric acid, in phosphate buffer 0.1M. Following postfixation in the same fixative overnight, the brains were cryoprotected in sucrose 30% overnight. Brains were then cut using a cryostat and the immunostainings were performed using the ‘free floating’ technique.