Production and characterization of monoclonal antibodies specific to lactotriaosylceramide.
We have established hybridoma cell lines producing monoclonal antibodies (mAbs) directed to N-acetylglucosaminylβ1-3galactose (GlcNAcβ1-3Gal) residue by immunizing BALB/c mice with lactotriaosylceramide (Lc(3)Cer). These obtained hybridoma cells, specific to Lc(3)Cer, were dual immunoglobulin (Ig)-producing cells which secreted both IgM and IgG molecules as antibodies. The established mAbs are able to react with not only Lc(3)Cer but also GlcNAcβ1-3-terminal glycosphingolipids (GSLs) despite branching or lactosamine chain lengths and human transferrin with terminal GlcNAc residues. Comparison of the variable regions of the cloned IgM and IgG by reversed transcription-polymerase chain reaction analysis confirmed that the variable regions determine the specificity, the other amino acids are conserved, and these mAbs are encoded by J558 and Vκ-21family genes. Furthermore, we have analyzed the expression of GSLs with GlcNAcβ1-3 epitope in acute leukemia cell lines and mouse fetal tissues using these mAbs, in which antigens were distributed comparatively. These mAbs are useful for studying the precise distribution of GlcNAcβ1-3Gal-terminating GSL expression in tissues as well as for detecting GSLs carrying terminal GlcNAcβ1-3Gal carbohydrate structure.
Nozaki
Hirofumi
H
Department of Agriculture and Life Science, Obihiro University of Agriculture and Veterinary Medicine, Obihiro, Hokkaido 080-8555, Japan.
Yanagida
Mayumi
M
Koide
Ken-ichi
K
Shiotani
Kazusa
K
Kinoshita
Mikio
M
Kobayashi
Yoshiyasu
Y
Watarai
Shinobu
S
Nakamura
Kazuo
K
Suzuki
Akemi
A
Ariga
Toshio
T
Kushi
Yasunori
Y
D000818:Animals; D058846:Antibodies, Monoclonal, Murine-Derived / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology*; D000918:Antibody Specificity / Q000235:genetics / Q000276:immunology*; D005260:Female; D018922:HL-60 Cells; D006801:Humans; D007074:Immunoglobulin G / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology*; D007075:Immunoglobulin M / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology*; D007135:Immunoglobulin Variable Region / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology*; D020014:K562 Cells; D007790:Lactosylceramides / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology*; D051379:Mice; D008807:Mice, Inbred BALB C; D020298:U937 Cells
D058846:Antibodies, Monoclonal, Murine-Derived; D007074:Immunoglobulin G; D007075:Immunoglobulin M; D007135:Immunoglobulin Variable Region; D007790:Lactosylceramides; C025575:lactotriaosylceramide