title: A novel monoclonal antibody against the C-terminal region of aquaporin-4.
abstract: Aquaporin-4 (AQP4), the most abundant water channel in the brain, plays a central role in water homeostasis, neuronal activity, and migration of astrocytes in the central nervous system. Recent studies have demonstrated that AQP4 is a target of an autoantibody specifically detected in an autoimmune neurologic disease called neuromyelitis optica. Here we have generated a monoclonal antibody (MAb) against the C-terminal region of AQP4 using a baculovirus expressing mouse AQP4 as an immunogen. This antibody (clone E5206) recognized both human and mouse AQP4s in a denaturing condition and was able to precipitate AQP4 from cell lysates of CHO cells stably expressing AQP4. Western blot analysis using deletion mutants revealed that the epitope was located within a region between Asp(303) and Leu(320) in the C-terminal tail of AQP4. Although clone E5206 could not be used for immunostaining when cells or tissues were fixed with 4% paraformaldehyde or 10% formalin, it could be used when cells were fixed with 10% trichloroacetic acid and when a formalin-fixed tissue section was pretreated with antigen-retrieval reagents. This MAb can be a valuable tool for analysis of AQP4 in a variety of physiological and pathophysiological contexts, in human tissues and organs as well as in rodent models, both in vitro and in vivo.
authors.affiliation: Department of Pharmacology, School of Medicine, Keio University, Shinjyuku-ku, Tokyo, Japan.
authors.forename: Julia
authors.initials: J
authors.lastname: Ramadhanti
authors.forename: Ping
authors.initials: P
authors.lastname: Huang
authors.forename: Osamu
authors.initials: O
authors.lastname: Kusano-Arai
authors.forename: Hiroko
authors.initials: H
authors.lastname: Iwanari
authors.forename: Toshiko
authors.initials: T
authors.lastname: Sakihama
authors.forename: Tasturo
authors.initials: T
authors.lastname: Misu
authors.forename: Kazuo
authors.initials: K
authors.lastname: Fujihara
authors.forename: Takao
authors.initials: T
authors.lastname: Hamakubo
authors.forename: Masato
authors.initials: M
authors.lastname: Yasui
authors.forename: Yoichiro
authors.initials: Y
authors.lastname: Abe
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000096:biosynthesis / Q000276:immunology* / Q000302:isolation & purification; D000917:Antibody Formation; D000918:Antibody Specificity; D051401:Aquaporin 4 / Q000276:immunology*; D015153:Blotting, Western; D001921:Brain / Q000276:immunology / Q000378:metabolism*; D016466:CHO Cells; D002478:Cells, Cultured; D006224:Cricetinae; D003412:Cricetulus; D004797:Enzyme-Linked Immunosorbent Assay; D005260:Female; D005455:Fluorescent Antibody Technique; D006801:Humans; D006825:Hybridomas / Q000276:immunology*; D007124:Immunoenzyme Techniques; D047468:Immunoprecipitation; D051379:Mice; D008807:Mice, Inbred BALB C; D008822:Mice, Transgenic; D008969:Molecular Sequence Data; D011994:Recombinant Proteins; D017386:Sequence Homology, Amino Acid
chemical_list: D000911:Antibodies, Monoclonal; D051401:Aquaporin 4; D011994:Recombinant Proteins
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