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Glycolipid‐directed FH6 monoclonal antibody recognizes high molecular weight glycprotein antigen carrying sialyl Le X ‐i determinant in the culture supernatant of PC‐9 cells
Author(s) -
Yamagata Yoko,
Shigeta Katsuyoshi,
Murachi Takashi,
Kannagi Reiji,
Masuda Tohru,
Imura Hiroo,
Shin Sadahito,
Tachikawa Tetsuya,
Noda Atsunari,
Ohe Yasuo
Publication year - 1988
Publication title -
journal of molecular recognition
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.401
H-Index - 79
eISSN - 1099-1352
pISSN - 0952-3499
DOI - 10.1002/jmr.300010303
Subject(s) - antigen , glycolipid , monoclonal antibody , microbiology and biotechnology , chemistry , radioimmunoassay , glycoprotein , biochemistry , antibody , antiserum , biology , immunology
The properties of the antigen recognized by monoclonal antibody FH6 have been analyzed. FH6 was originally generated against a glycolipid, i.e. a difucoganglioside isolated from human colonic adenocarcinoma, and specifically reacts with sialyl Le x ‐i detreminant. Several culture supernatants of human carcinoma cell line cells were found to have high levels of FH6‐reactive antigen, and PC‐9, a human lung carcinoma cell line was used for the analysis. A solid‐phase sandwich radioimmunoassay was performed to detect the antigen. The antigenic activity was extractable in 0.6 M PCA or 7% TCA, and was sensitive to mild alkaline treatment and to Pronase digestion. Most of the antigen was eluted in the void volume of a Sepharose CL‐2B column, which indicates that its molecular weights is greater than several million. It was eluted from a DEAE‐cellulose column at a NaCl concentration in the range of 0.2–0.25 M . The immunoaffinity‐purified antigen has a high carbohydrate content of more than 80%. These data indicate that the antigen recognized by FH6 in the culture supernatant of PC‐9 is not a glycolipid, but a high molecular weight glycoprotein which could be referred to as a mucin, or a proteoglycan, which contains keratan‐sulfate like glycosaminoglycan chains, as judged from the results of the glycosidase treatments.