Epitope mapping of an anti-alpha thalassemia/mental retardation syndrome X-linked monoclonal antibody AMab-6

The alpha-thalassemia/mental-retardation-syndrome-X-linked (ATRX) gene is located on the q arm of the X chromosome. ATRX gene mutations were first discovered in pancreatic neuroendocrine tumors, and subsequently in other cancer subtypes, including gliomas. Molecular subgrouping of gliomas has been m...

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Published inBiochemistry and biophysics reports Vol. 15; pp. 76 - 80
Main Authors Kaneko, Mika K, Yamada, Shinji, Itai, Shunsuke, Furusawa, Yoshikazu, Nakamura, Takuro, Yanaka, Miyuki, Handa, Saori, Hisamatsu, Kayo, Nakamura, Yoshimi, Fukui, Masato, Harada, Hiroyuki, Kato, Yukinari
Format Journal Article
LanguageEnglish
Published Netherlands Elsevier 01.09.2018
Subjects
ATRX, alpha-thalassemia/mental-retardation-syndrome-X-linked
AMab-6
PBS, phosphate-buffered saline
mAb, monoclonal antibody
ATRX
DAB, 3,3-diaminobenzidine tetrahydrochloride
Epitope mapping
ELISA, enzyme-linked immunosorbent assay
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Abstract The alpha-thalassemia/mental-retardation-syndrome-X-linked (ATRX) gene is located on the q arm of the X chromosome. ATRX gene mutations were first discovered in pancreatic neuroendocrine tumors, and subsequently in other cancer subtypes, including gliomas. Molecular subgrouping of gliomas has been more important than conventional histological classifications. Mutations in the isocitrate dehydrogenase (IDH), telomerase reverse transcriptase (TERT) promoter, and ATRX and the codeletion of chromosomes 1p/19q are used as biomarkers for diagnosing the subtypes of diffuse gliomas. We recently developed a sensitive monoclonal antibody (mAb) AMab-6 against ATRX by immunizing mice with recombinant human ATRX. AMab-6 can help to detect ATRX mutations via Western blotting and immunohistochemical analyses. In this study, we characterized the binding epitope of AMab-6 using enzyme-linked immunosorbent assay (ELISA), Western blotting, and immunohistochemical analysis, and found that Gln2368 of ATRX is critical for AMab-6 binding to ATRX. Our findings could be applied to the production of more functional anti-ATRX mAbs.
AbstractList The alpha-thalassemia/mental-retardation-syndrome-X-linked (ATRX) gene is located on the q arm of the X chromosome. ATRX gene mutations were first discovered in pancreatic neuroendocrine tumors, and subsequently in other cancer subtypes, including gliomas. Molecular subgrouping of gliomas has been more important than conventional histological classifications. Mutations in the isocitrate dehydrogenase (IDH), telomerase reverse transcriptase (TERT) promoter, and ATRX and the codeletion of chromosomes 1p/19q are used as biomarkers for diagnosing the subtypes of diffuse gliomas. We recently developed a sensitive monoclonal antibody (mAb) AMab-6 against ATRX by immunizing mice with recombinant human ATRX. AMab-6 can help to detect ATRX mutations via Western blotting and immunohistochemical analyses. In this study, we characterized the binding epitope of AMab-6 using enzyme-linked immunosorbent assay (ELISA), Western blotting, and immunohistochemical analysis, and found that Gln2368 of ATRX is critical for AMab-6 binding to ATRX. Our findings could be applied to the production of more functional anti-ATRX mAbs. • ATRX gene is located on the q arm of the X chromosome. • AMab-6 detected ATRX in Western blot and immunohistochemical analyses sensitively. • Gln2368 of ATRX is critical for AMab-6-binding to ATRX. • A blocking peptide completely neutralized AMab-6 reaction by IHC.
The alpha-thalassemia/mental-retardation-syndrome-X-linked (ATRX) gene is located on the q arm of the X chromosome. ATRX gene mutations were first discovered in pancreatic neuroendocrine tumors, and subsequently in other cancer subtypes, including gliomas. Molecular subgrouping of gliomas has been more important than conventional histological classifications. Mutations in the isocitrate dehydrogenase (IDH), telomerase reverse transcriptase (TERT) promoter, and ATRX and the codeletion of chromosomes 1p/19q are used as biomarkers for diagnosing the subtypes of diffuse gliomas. We recently developed a sensitive monoclonal antibody (mAb) AMab-6 against ATRX by immunizing mice with recombinant human ATRX. AMab-6 can help to detect ATRX mutations via Western blotting and immunohistochemical analyses. In this study, we characterized the binding epitope of AMab-6 using enzyme-linked immunosorbent assay (ELISA), Western blotting, and immunohistochemical analysis, and found that Gln2368 of ATRX is critical for AMab-6 binding to ATRX. Our findings could be applied to the production of more functional anti-ATRX mAbs. Keywords: ATRX, AMab-6, Epitope mapping
The alpha-thalassemia/mental-retardation-syndrome-X-linked (ATRX) gene is located on the q arm of the X chromosome. ATRX gene mutations were first discovered in pancreatic neuroendocrine tumors, and subsequently in other cancer subtypes, including gliomas. Molecular subgrouping of gliomas has been more important than conventional histological classifications. Mutations in the isocitrate dehydrogenase (IDH), telomerase reverse transcriptase (TERT) promoter, and ATRX and the codeletion of chromosomes 1p/19q are used as biomarkers for diagnosing the subtypes of diffuse gliomas. We recently developed a sensitive monoclonal antibody (mAb) AMab-6 against ATRX by immunizing mice with recombinant human ATRX. AMab-6 can help to detect ATRX mutations via Western blotting and immunohistochemical analyses. In this study, we characterized the binding epitope of AMab-6 using enzyme-linked immunosorbent assay (ELISA), Western blotting, and immunohistochemical analysis, and found that Gln2368 of ATRX is critical for AMab-6 binding to ATRX. Our findings could be applied to the production of more functional anti-ATRX mAbs.
Author Kato, Yukinari
Yamada, Shinji
Handa, Saori
Kaneko, Mika K
Yanaka, Miyuki
Hisamatsu, Kayo
Nakamura, Yoshimi
Fukui, Masato
Nakamura, Takuro
Itai, Shunsuke
Furusawa, Yoshikazu
Harada, Hiroyuki
AuthorAffiliation c New Industry Creation Hatchery Center, Tohoku University, 2-1, Seiryo-machi, Aoba-ku, Sendai, Miyagi 980-8575, Japan
d ZENOAQ RESOURCE CO., LTD., 1-1 Tairanoue, Sasagawa, Asaka-machi, Koriyama, Fukushima 963-0196, Japan
a Department of Antibody Drug Development, Tohoku University Graduate School of Medicine, 2-1 Seiryo-machi, Aoba-ku, Sendai, Miyagi 980-8575, Japan
b Department of Oral and Maxillofacial Surgery, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University, 1-5-45, Yushima, Bunkyo-ku, Tokyo 113-8510, Japan
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  surname: Handa
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Cites_doi 10.1007/s00401-015-1409-0
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Keywords ATRX, alpha-thalassemia/mental-retardation-syndrome-X-linked
AMab-6
PBS, phosphate-buffered saline
mAb, monoclonal antibody
ATRX
DAB, 3,3-diaminobenzidine tetrahydrochloride
Epitope mapping
ELISA, enzyme-linked immunosorbent assay
Language English
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PublicationTitle Biochemistry and biophysics reports
PublicationTitleAlternate Biochem Biophys Rep
PublicationYear 2018
Publisher Elsevier
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Snippet The alpha-thalassemia/mental-retardation-syndrome-X-linked (ATRX) gene is located on the q arm of the X chromosome. ATRX gene mutations were first discovered...
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Title Epitope mapping of an anti-alpha thalassemia/mental retardation syndrome X-linked monoclonal antibody AMab-6
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