title: In situ visualization of a glycoform of transferrin: localization of α2,6-sialylated transferrin in the liver.
abstract: We previously found that a lectin, Sambucus sieboldiana agglutinin (SSA), bound to α2,6-sialylated glycan epitopes on transferrin and inhibited anti-transferrin antibody binding to the antigen in ELISA (SSA inhibition). Here we report that SSA inhibition is applicable to immunohistochemistry, localizing α2,6-sialylated transferrin in the liver. Immunohistochemistry using anti-transferrin polyclonal antibody revealed that transferrin was detected in hepatocytes near interlobular veins. Addition of SSA lectin markedly attenuated the staining. Sialidase treatment of a liver section abolished SSA binding and concomitantly cancelled SSA inhibition, suggesting that SSA binding to glycan epitopes on the section was essential for the inhibition. To examine the importance of proximity between antigen epitopes and SSA-binding (glycosylation) sites, we prepared two anti-peptide antibodies against partial amino acid sequences of transferrin. One antibody (Tf-596Ab) is against a peptide sequence, Cys596-Ala614, which is proximal to N-glycosylation sites (Asn-432 and Asn-630). The other (Tf-120Ab) is against a peptide sequence, Val120-Cys137, distal to the sites. The staining signals of Tf-596Ab were reduced by the addition of SSA, whereas those of Tf-120Ab were reduced only a little. This result suggests that proximity of the antigen epitope to SSA binding sites is critical for SSA inhibition in immunohistochemistry.
authors.affiliation: Department of Neurosurgery and Department of Biochemistry, Fukushima Medical University, 1 Hikarigaoka, Fukushima, Fukushima 960-1295, Japan; Structural Glycobiology Team, Systems Glycobiology Research Group, RIKEN-Max Planck Joint Research Center for Systems Chemical Biology, Global Research Cluster, RIKEN, 2-1 Hirosawa, Wako-Shi, Saitama 351-0198, Japan; Department of Biological Sciences, Immuno-Biological Laboratories, Co. Ltd., 1091-1 Naka, Fujioka, Gunma 375-0005, Japan; GeneticLab Co., Ltd., 15-28-196 Kita 9-Jyu, Nishi, Tyuou-Ku, Sapporo-City, Hokkaido 060-009, Japan; Department of Cell Science, Department of Human Life Science; and Fukushima Industry-University-Government Research Center, Fukushima Medical University, 1 Hikarigaoka, Fukushima, Fukushima 960-1295, Japan yasuc@fmu.ac.jp ponchan@fmu.ac.jp.
authors.forename: Yuka
authors.initials: Y
authors.lastname: Matsumoto
authors.affiliation: Department of Neurosurgery and Department of Biochemistry, Fukushima Medical University, 1 Hikarigaoka, Fukushima, Fukushima 960-1295, Japan; Structural Glycobiology Team, Systems Glycobiology Research Group, RIKEN-Max Planck Joint Research Center for Systems Chemical Biology, Global Research Cluster, RIKEN, 2-1 Hirosawa, Wako-Shi, Saitama 351-0198, Japan; Department of Biological Sciences, Immuno-Biological Laboratories, Co. Ltd., 1091-1 Naka, Fujioka, Gunma 375-0005, Japan; GeneticLab Co., Ltd., 15-28-196 Kita 9-Jyu, Nishi, Tyuou-Ku, Sapporo-City, Hokkaido 060-009, Japan; Department of Cell Science, Department of Human Life Science; and Fukushima Industry-University-Government Research Center, Fukushima Medical University, 1 Hikarigaoka, Fukushima, Fukushima 960-1295, Japan yasuc@fmu.ac.jp ponchan@fmu.ac.jp.
authors.forename: Toshie
authors.initials: T
authors.lastname: Saito
authors.affiliation: Department of Neurosurgery and Department of Biochemistry, Fukushima Medical University, 1 Hikarigaoka, Fukushima, Fukushima 960-1295, Japan; Structural Glycobiology Team, Systems Glycobiology Research Group, RIKEN-Max Planck Joint Research Center for Systems Chemical Biology, Global Research Cluster, RIKEN, 2-1 Hirosawa, Wako-Shi, Saitama 351-0198, Japan; Department of Biological Sciences, Immuno-Biological Laboratories, Co. Ltd., 1091-1 Naka, Fujioka, Gunma 375-0005, Japan; GeneticLab Co., Ltd., 15-28-196 Kita 9-Jyu, Nishi, Tyuou-Ku, Sapporo-City, Hokkaido 060-009, Japan; Department of Cell Science, Department of Human Life Science; and Fukushima Industry-University-Government Research Center, Fukushima Medical University, 1 Hikarigaoka, Fukushima, Fukushima 960-1295, Japan yasuc@fmu.ac.jp ponchan@fmu.ac.jp.
authors.forename: Kyoka
authors.initials: K
authors.lastname: Hoshi
authors.affiliation: Department of Neurosurgery and Department of Biochemistry, Fukushima Medical University, 1 Hikarigaoka, Fukushima, Fukushima 960-1295, Japan; Structural Glycobiology Team, Systems Glycobiology Research Group, RIKEN-Max Planck Joint Research Center for Systems Chemical Biology, Global Research Cluster, RIKEN, 2-1 Hirosawa, Wako-Shi, Saitama 351-0198, Japan; Department of Biological Sciences, Immuno-Biological Laboratories, Co. Ltd., 1091-1 Naka, Fujioka, Gunma 375-0005, Japan; GeneticLab Co., Ltd., 15-28-196 Kita 9-Jyu, Nishi, Tyuou-Ku, Sapporo-City, Hokkaido 060-009, Japan; Department of Cell Science, Department of Human Life Science; and Fukushima Industry-University-Government Research Center, Fukushima Medical University, 1 Hikarigaoka, Fukushima, Fukushima 960-1295, Japan.
authors.forename: Hiromi
authors.initials: H
authors.lastname: Ito
authors.affiliation: Department of Neurosurgery and Department of Biochemistry, Fukushima Medical University, 1 Hikarigaoka, Fukushima, Fukushima 960-1295, Japan; Structural Glycobiology Team, Systems Glycobiology Research Group, RIKEN-Max Planck Joint Research Center for Systems Chemical Biology, Global Research Cluster, RIKEN, 2-1 Hirosawa, Wako-Shi, Saitama 351-0198, Japan; Department of Biological Sciences, Immuno-Biological Laboratories, Co. Ltd., 1091-1 Naka, Fujioka, Gunma 375-0005, Japan; GeneticLab Co., Ltd., 15-28-196 Kita 9-Jyu, Nishi, Tyuou-Ku, Sapporo-City, Hokkaido 060-009, Japan; Department of Cell Science, Department of Human Life Science; and Fukushima Industry-University-Government Research Center, Fukushima Medical University, 1 Hikarigaoka, Fukushima, Fukushima 960-1295, Japan.
authors.forename: Yoshinobu
authors.initials: Y
authors.lastname: Kariya
authors.affiliation: Department of Neurosurgery and Department of Biochemistry, Fukushima Medical University, 1 Hikarigaoka, Fukushima, Fukushima 960-1295, Japan; Structural Glycobiology Team, Systems Glycobiology Research Group, RIKEN-Max Planck Joint Research Center for Systems Chemical Biology, Global Research Cluster, RIKEN, 2-1 Hirosawa, Wako-Shi, Saitama 351-0198, Japan; Department of Biological Sciences, Immuno-Biological Laboratories, Co. Ltd., 1091-1 Naka, Fujioka, Gunma 375-0005, Japan; GeneticLab Co., Ltd., 15-28-196 Kita 9-Jyu, Nishi, Tyuou-Ku, Sapporo-City, Hokkaido 060-009, Japan; Department of Cell Science, Department of Human Life Science; and Fukushima Industry-University-Government Research Center, Fukushima Medical University, 1 Hikarigaoka, Fukushima, Fukushima 960-1295, Japan.
authors.forename: Masamichi
authors.initials: M
authors.lastname: Nagae
authors.affiliation: Department of Neurosurgery and Department of Biochemistry, Fukushima Medical University, 1 Hikarigaoka, Fukushima, Fukushima 960-1295, Japan; Structural Glycobiology Team, Systems Glycobiology Research Group, RIKEN-Max Planck Joint Research Center for Systems Chemical Biology, Global Research Cluster, RIKEN, 2-1 Hirosawa, Wako-Shi, Saitama 351-0198, Japan; Department of Biological Sciences, Immuno-Biological Laboratories, Co. Ltd., 1091-1 Naka, Fujioka, Gunma 375-0005, Japan; GeneticLab Co., Ltd., 15-28-196 Kita 9-Jyu, Nishi, Tyuou-Ku, Sapporo-City, Hokkaido 060-009, Japan; Department of Cell Science, Department of Human Life Science; and Fukushima Industry-University-Government Research Center, Fukushima Medical University, 1 Hikarigaoka, Fukushima, Fukushima 960-1295, Japan.
authors.forename: Yoshiki
authors.initials: Y
authors.lastname: Yamaguchi
authors.affiliation: Department of Neurosurgery and Department of Biochemistry, Fukushima Medical University, 1 Hikarigaoka, Fukushima, Fukushima 960-1295, Japan; Structural Glycobiology Team, Systems Glycobiology Research Group, RIKEN-Max Planck Joint Research Center for Systems Chemical Biology, Global Research Cluster, RIKEN, 2-1 Hirosawa, Wako-Shi, Saitama 351-0198, Japan; Department of Biological Sciences, Immuno-Biological Laboratories, Co. Ltd., 1091-1 Naka, Fujioka, Gunma 375-0005, Japan; GeneticLab Co., Ltd., 15-28-196 Kita 9-Jyu, Nishi, Tyuou-Ku, Sapporo-City, Hokkaido 060-009, Japan; Department of Cell Science, Department of Human Life Science; and Fukushima Industry-University-Government Research Center, Fukushima Medical University, 1 Hikarigaoka, Fukushima, Fukushima 960-1295, Japan.
authors.forename: Yoshiaki
authors.initials: Y
authors.lastname: Hagiwara
authors.affiliation: Department of Neurosurgery and Department of Biochemistry, Fukushima Medical University, 1 Hikarigaoka, Fukushima, Fukushima 960-1295, Japan; Structural Glycobiology Team, Systems Glycobiology Research Group, RIKEN-Max Planck Joint Research Center for Systems Chemical Biology, Global Research Cluster, RIKEN, 2-1 Hirosawa, Wako-Shi, Saitama 351-0198, Japan; Department of Biological Sciences, Immuno-Biological Laboratories, Co. Ltd., 1091-1 Naka, Fujioka, Gunma 375-0005, Japan; GeneticLab Co., Ltd., 15-28-196 Kita 9-Jyu, Nishi, Tyuou-Ku, Sapporo-City, Hokkaido 060-009, Japan; Department of Cell Science, Department of Human Life Science; and Fukushima Industry-University-Government Research Center, Fukushima Medical University, 1 Hikarigaoka, Fukushima, Fukushima 960-1295, Japan.
authors.forename: Noriaki
authors.initials: N
authors.lastname: Kinoshita
authors.affiliation: Department of Neurosurgery and Department of Biochemistry, Fukushima Medical University, 1 Hikarigaoka, Fukushima, Fukushima 960-1295, Japan; Structural Glycobiology Team, Systems Glycobiology Research Group, RIKEN-Max Planck Joint Research Center for Systems Chemical Biology, Global Research Cluster, RIKEN, 2-1 Hirosawa, Wako-Shi, Saitama 351-0198, Japan; Department of Biological Sciences, Immuno-Biological Laboratories, Co. Ltd., 1091-1 Naka, Fujioka, Gunma 375-0005, Japan; GeneticLab Co., Ltd., 15-28-196 Kita 9-Jyu, Nishi, Tyuou-Ku, Sapporo-City, Hokkaido 060-009, Japan; Department of Cell Science, Department of Human Life Science; and Fukushima Industry-University-Government Research Center, Fukushima Medical University, 1 Hikarigaoka, Fukushima, Fukushima 960-1295, Japan.
authors.forename: Ikuo
authors.initials: I
authors.lastname: Wada
authors.affiliation: Department of Neurosurgery and Department of Biochemistry, Fukushima Medical University, 1 Hikarigaoka, Fukushima, Fukushima 960-1295, Japan; Structural Glycobiology Team, Systems Glycobiology Research Group, RIKEN-Max Planck Joint Research Center for Systems Chemical Biology, Global Research Cluster, RIKEN, 2-1 Hirosawa, Wako-Shi, Saitama 351-0198, Japan; Department of Biological Sciences, Immuno-Biological Laboratories, Co. Ltd., 1091-1 Naka, Fujioka, Gunma 375-0005, Japan; GeneticLab Co., Ltd., 15-28-196 Kita 9-Jyu, Nishi, Tyuou-Ku, Sapporo-City, Hokkaido 060-009, Japan; Department of Cell Science, Department of Human Life Science; and Fukushima Industry-University-Government Research Center, Fukushima Medical University, 1 Hikarigaoka, Fukushima, Fukushima 960-1295, Japan.
authors.forename: Kiyoshi
authors.initials: K
authors.lastname: Saito
authors.affiliation: Department of Neurosurgery and Department of Biochemistry, Fukushima Medical University, 1 Hikarigaoka, Fukushima, Fukushima 960-1295, Japan; Structural Glycobiology Team, Systems Glycobiology Research Group, RIKEN-Max Planck Joint Research Center for Systems Chemical Biology, Global Research Cluster, RIKEN, 2-1 Hirosawa, Wako-Shi, Saitama 351-0198, Japan; Department of Biological Sciences, Immuno-Biological Laboratories, Co. Ltd., 1091-1 Naka, Fujioka, Gunma 375-0005, Japan; GeneticLab Co., Ltd., 15-28-196 Kita 9-Jyu, Nishi, Tyuou-Ku, Sapporo-City, Hokkaido 060-009, Japan; Department of Cell Science, Department of Human Life Science; and Fukushima Industry-University-Government Research Center, Fukushima Medical University, 1 Hikarigaoka, Fukushima, Fukushima 960-1295, Japan Department of Neurosurgery and Department of Biochemistry, Fukushima Medical University, 1 Hikarigaoka, Fukushima, Fukushima 960-1295, Japan; Structural Glycobiology Team, Systems Glycobiology Research Group, RIKEN-Max Planck Joint Research Center for Systems Chemical Biology, Global Research Cluster, RIKEN, 2-1 Hirosawa, Wako-Shi, Saitama 351-0198, Japan; Department of Biological Sciences, Immuno-Biological Laboratories, Co. Ltd., 1091-1 Naka, Fujioka, Gunma 375-0005, Japan; GeneticLab Co., Ltd., 15-28-196 Kita 9-Jyu, Nishi, Tyuou-Ku, Sapporo-City, Hokkaido 060-009, Japan; Department of Cell Science, Department of Human Life Science; and Fukushima Industry-University-Government Research Center, Fukushima Medical University, 1 Hikarigaoka, Fukushima, Fukushima 960-1295, Japan yasuc@fmu.ac.jp.
authors.forename: Takashi
authors.initials: T
authors.lastname: Honda
authors.affiliation: Department of Neurosurgery and Department of Biochemistry, Fukushima Medical University, 1 Hikarigaoka, Fukushima, Fukushima 960-1295, Japan; Structural Glycobiology Team, Systems Glycobiology Research Group, RIKEN-Max Planck Joint Research Center for Systems Chemical Biology, Global Research Cluster, RIKEN, 2-1 Hirosawa, Wako-Shi, Saitama 351-0198, Japan; Department of Biological Sciences, Immuno-Biological Laboratories, Co. Ltd., 1091-1 Naka, Fujioka, Gunma 375-0005, Japan; GeneticLab Co., Ltd., 15-28-196 Kita 9-Jyu, Nishi, Tyuou-Ku, Sapporo-City, Hokkaido 060-009, Japan; Department of Cell Science, Department of Human Life Science; and Fukushima Industry-University-Government Research Center, Fukushima Medical University, 1 Hikarigaoka, Fukushima, Fukushima 960-1295, Japan Department of Neurosurgery and Department of Biochemistry, Fukushima Medical University, 1 Hikarigaoka, Fukushima, Fukushima 960-1295, Japan; Structural Glycobiology Team, Systems Glycobiology Research Group, RIKEN-Max Planck Joint Research Center for Systems Chemical Biology, Global Research Cluster, RIKEN, 2-1 Hirosawa, Wako-Shi, Saitama 351-0198, Japan; Department of Biological Sciences, Immuno-Biological Laboratories, Co. Ltd., 1091-1 Naka, Fujioka, Gunma 375-0005, Japan; GeneticLab Co., Ltd., 15-28-196 Kita 9-Jyu, Nishi, Tyuou-Ku, Sapporo-City, Hokkaido 060-009, Japan; Department of Cell Science, Department of Human Life Science; and Fukushima Industry-University-Government Research Center, Fukushima Medical University, 1 Hikarigaoka, Fukushima, Fukushima 960-1295, Japan ponchan@fmu.ac.jp.
authors.forename: Yasuhiro
authors.initials: Y
authors.lastname: Hashimoto
mesh_terms: D000595:Amino Acid Sequence; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D015151:Immunoblotting; D037102:Lectins / Q000378:metabolism; D008099:Liver / Q000378:metabolism*; D008969:Molecular Sequence Data; D009439:Neuraminidase / Q000737:chemistry / Q000378:metabolism; D020033:Protein Isoforms / Q000378:metabolism; D021381:Protein Transport; D014168:Transferrin / Q000378:metabolism*
chemical_list: D037102:Lectins; D020033:Protein Isoforms; D014168:Transferrin; D009439:Neuraminidase
keywords: Sambucus sieboldiana agglutinin (SSA) lectin; immunohistochemistry; transferrin; α2,6-sialic acid