Establishment of an antigen-specific B cell clone by somatic hybridization.
Splenic B cells of A/J mice immunized with 2,4,6-trinitrophenyl (TNP)-lipopolysaccharide were fused with 2.52M, a mutant of a B cell line, in the presence of polyethylene glycol and dimethyl sulfoxide. TP67.21, a subclone of a resulting hybridoma, expresses IAk, IEk, IgM, B220, P50, and receptors for C3 fragment of complement, the Fc portion of IgG, and interleukin 2 receptor on the cell membrane; it also possesses receptor molecules for TNP on its surface, derived from TNP-reactive B cells of A/J mice primed with TNP-lipopolysaccharide used for somatic hybridization, by a rosette-forming assay with TNP-sheep erythrocytes. In contrast, parental 2.52M lacks IAk and IEk on the cell membrane and does not bind to TNP-sheep erythrocytes under the same conditions. Thus, it is likely that TP67.21 is an antigen-specific B cell clone directed against TNP. The antigen binding of cells was markedly inhibited by the specific free hapten or anti-IgM antibodies. Interestingly, TP67.21 was induced to generate a significant amount of anti-TNP antibody when treated with TNP conjugates including T cell-independent and -dependent antigens, such as TNP-lipopolysaccharide, TNP-bovine serum albumin, TNP-ovalbumin, and TNP-keyhole limpet hemocyanine in the absence of T cell help, as well as polyclonal activators; this was followed by a marked decrease in the expression of B cell surface markers on the cell membrane. This suggests that the cross-linkage of receptor molecules on TP67.21 by antigen may directly provide a differentiative signal for maturation to a lineage of B cells, and consequently results in the generation of antigen-specific antibodies without T cell involvement.
Hamano
T
T
Second Department of Internal Medicine, Hyogo College of Medicine, Japan.
Murata
Y
Y
Yamasaki
T
T
Yasuda
Y
Y
Iwasaki
T
T
Nagai
K
K
D000818:Animals; D000917:Antibody Formation; D000941:Antigens / Q000276:immunology; D000954:Antigens, Surface / Q000032:analysis; D001402:B-Lymphocytes / Q000166:cytology* / Q000276:immunology; D002454:Cell Differentiation; D002459:Cell Fusion; D000949:Histocompatibility Antigens Class II / Q000032:analysis; D008213:Lymphocyte Activation; D051379:Mice; D011947:Receptors, Antigen, B-Cell / Q000032:analysis; D011956:Receptors, Cell Surface / Q000276:immunology; D012397:Rosette Formation; D014301:Trinitrobenzenes / Q000276:immunology
D000941:Antigens; D000954:Antigens, Surface; D000949:Histocompatibility Antigens Class II; D011947:Receptors, Antigen, B-Cell; D011956:Receptors, Cell Surface; D014301:Trinitrobenzenes