title: Methods for immunoblot detection and electron microscopic localization of septin subunits in mammalian nervous systems.
abstract: The minimal functional units of the mammalian septin system are diverse heterooligomers of SEPT1-14 subunits, which are most abundantly and differentially expressed in postmitotic neurons and glia. The subunit compositions of such heterooligomers are thought to differentiate their affinity for other proteins and lipids, and subcellular localization. Thus, high-precision quantification and mapping of each subunit is necessary to understand their subcellular functions and physiological roles. However, systematic information on the localization of individual septin subunits in the mammalian nervous system is limited. Here, we present our experimental workflows for the study of septin expression and localization in the rodent brain by immunoblot and serial section immunoelectron microscopy. Our protocols, based on standard methods, have been rigorously optimized and simplified for universality and reproducibility to aid non-experts in the field.
authors.affiliation: University of Tokyo, Tokyo, Japan.
authors.forename: L K
authors.initials: LK
authors.lastname: Parajuli
authors.affiliation: Nagoya University Graduate School of Science, Nagoya, Japan.
authors.forename: N
authors.initials: N
authors.lastname: Ageta-Ishihara
authors.affiliation: Fujita Health University, Toyoake, Japan.
authors.forename: H
authors.initials: H
authors.lastname: Ageta
authors.affiliation: University of Fukui, Fukui, Japan.
authors.forename: Y
authors.initials: Y
authors.lastname: Fukazawa
authors.affiliation: Nagoya University Graduate School of Science, Nagoya, Japan.
authors.forename: M
authors.initials: M
authors.lastname: Kinoshita
mesh_terms: D000818:Animals; D015151:Immunoblotting / Q000379:methods*; D008322:Mammals; D051379:Mice; D008854:Microscopy, Electron / Q000379:methods*; D009420:Nervous System / Q000737:chemistry / Q000648:ultrastructure*; D009474:Neurons / Q000737:chemistry / Q000648:ultrastructure; D021122:Protein Subunits / Q000737:chemistry / Q000302:isolation & purification; D051381:Rats; D058112:Septins / Q000737:chemistry / Q000302:isolation & purification*
chemical_list: D021122:Protein Subunits; D058112:Septins
keywords: Electron microscopy; Immunoblot; Immunogold labeling; Nervous system; Proteins; Septins