Further analyses of epitopes for human monoclonal anti-basement membrane zone antibodies produced by stable human hybridoma cell lines constructed with epstein-barr virus transformants

Takashi Hashimoto, Masayuki Amagai, Tamotsu Ebihara, Shinobu Gamou, Nobuyoshi Shimizu, Takuichi Tsubata, Akira Hasegawa, Keizaburo Miki, Takeji Nishikawa

研究成果: Article査読

36 被引用数 (Scopus)

抄録

We previously established Epstein-Barr virus (EBV)-transformed bullous pemphigoid (BP) patient lymphoblastoid cell lines, which produced human monoclonal anti-basement membrane zone antibodies. In the present study, we established two independent human-human hybridomas by fusion of these EBV transformants with a human B-cell line. These hybridomas, designated 5E-HY-4B and 10D-HY-8B, were very stable and showed a high yield of monoclonal antibody (MoAb) secretion. Each cell line was tetraploid and showed combined rearranged segments of immunoglobulin heavy-chain gene derived from both an EBV transformant and a parent cell. Immunoblot analysis showed that the 5E-HY-4B MoAb recognized the 230-kDa BP antigen but that the 10D-HY-8B MoAb did not show any reactivity. In contrast, both MoAbs precipitated the 230-kDa BP antigen with immunoprecipitation. These results indicate that the two MoAbs reacted with different epitopes on the 230-kDa BP antigen: a continuous epitope for the 5E-HY-4B MoAb and a conformation-dependent epitope for the 10D-HY-8B MoAb. This speculation was confirmed at the molecular level by the result that the fusion protein produced by a partial cDNA for the 230-kDa mouse BP antigen reacted with the 5E-HY-4B MoAb but not with the 10D-HY-8B MoAb. Furthermore, the study of the reactivity with fusion proteins of a series of deleted clones restricted the epitope for the 5E-HY-4B MoAb within the region with 114 amino acid residues in the C-terminal domain of the 230-kDa BP antigen.

本文言語English
ページ(範囲)310-315
ページ数6
ジャーナルJournal of Investigative Dermatology
100
3
DOI
出版ステータスPublished - 1993 3月

ASJC Scopus subject areas

  • 生化学
  • 分子生物学
  • 皮膚病学
  • 細胞生物学

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