TY - JOUR
T1 - In vitro immunization of rat spleen lymphocytes with Forssman glycosphingolipid antigen and the generation of a monoclonal antibody
AU - Gathuru, J. K.
AU - Miyoshi, I.
AU - Naiki, M.
PY - 1991
Y1 - 1991
N2 - A procedure for in vitro immunization of splenic lymphocytes with a glycolipid antigen is described. Culture medium supernatant of ConA- and PHA-stimulated spleen cells and that of Con A-stimulated human Jurkat T cell line (IL-2-rich medium) were used as sources of cytokines to support T and B cell stimulation, and anti-μ was used to support B cell differentiation. Unprimed rat spleen cells (2 × 106/ml) were stimulated with 2 μg/ml Forssman glycolipid antigen coupled to Sepharose for 4 days. The cells were fused with a mouse myeloma cell line P3-X63-Ag8-U1. At initial screening, 12% of the colony forming wells were secreting specific antibody. After cloning, a stable hybridoma cell line (designated 4C3) was established which secreted a monoclonal IgM antibody directed against the carbohydrate moiety of Forssman glycosphingolipid (GalNAcα1-3GalNAcβ1-3Galα1-4Galβ1-4Glc-ceramide).
AB - A procedure for in vitro immunization of splenic lymphocytes with a glycolipid antigen is described. Culture medium supernatant of ConA- and PHA-stimulated spleen cells and that of Con A-stimulated human Jurkat T cell line (IL-2-rich medium) were used as sources of cytokines to support T and B cell stimulation, and anti-μ was used to support B cell differentiation. Unprimed rat spleen cells (2 × 106/ml) were stimulated with 2 μg/ml Forssman glycolipid antigen coupled to Sepharose for 4 days. The cells were fused with a mouse myeloma cell line P3-X63-Ag8-U1. At initial screening, 12% of the colony forming wells were secreting specific antibody. After cloning, a stable hybridoma cell line (designated 4C3) was established which secreted a monoclonal IgM antibody directed against the carbohydrate moiety of Forssman glycosphingolipid (GalNAcα1-3GalNAcβ1-3Galα1-4Galβ1-4Glc-ceramide).
KW - Forssman glycosphingolipid
KW - In vitro immunization
KW - Monoclonal antibody
KW - Tumor-associated antigen
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U2 - 10.1016/0022-1759(91)90398-Y
DO - 10.1016/0022-1759(91)90398-Y
M3 - Article
C2 - 2010622
AN - SCOPUS:0026027193
SN - 0022-1759
VL - 137
SP - 95
EP - 102
JO - Journal of Immunological Methods
JF - Journal of Immunological Methods
IS - 1
ER -