Diversity in the available repertoire of murine antibodies reactive with bromelain-treated isologous erythrocytes.
Antibodies specific for bromelain-treated mouse RBC (BrMRBC) are of interest as models of "natural autoantibodies" and because of their primary source is Ly-1+ (CD5+) B cells. In earlier work by others, anti-BrMRBC hybridomas prepared by using CBA or NZB "spontaneously activating" peritoneal B cells were all found to produce mAb with a single common H chain V region sequence, by using a novel gene (VH11p), and a single common L chain V region sequence, a member of the Vk9 group (VkBrMp). We prepared anti-BrMRBC hybridomas by using LPS-activated B10.A splenic B cells in order to reveal the maximum available diversity in this repertoire. Data based on binding studies, Northern blot analyses with V region-specific probes, and mRNA nucleotide sequence analysis indicated that there is combining-site diversity in the repertoire of anti-BrMRBC hybridomas. There was considerable variation in trimethylammonium (a constituent of phosphatidyl choline) binding efficiency, and one of the anti-BrMRBC mAb showed no detectable binding. Northern blot analyses indicated 6 of 11 mAb to be of the VH11p/VkBrMp type, including one dual reactive anti-[BrMRBC + SRBC] mAb. Sequence analyses of the H chain V regions of four of the non-VH11p mAb revealed utilization of four distinct VH, three of which are very similar to the VH expressed by Ly-1+ B cell clones or lymphomas, as reported by others. However, because the VH11p/VkBrMp-type mAb were all relatively efficient at lysing BrMRBC and binding trimethylammonium, we suggest that affinity considerations may determine the selective predominance of B cells with this V region configuration from an available repertoire of considerable diversity.
Conger
J D
JD
Department of Microbiology and Immunology, Duke University Medical Center, Durham, NC 27710.
Sage
H J
HJ
Corley
R B
RB
D000818:Animals; D000911:Antibodies, Monoclonal / Q000235:genetics / Q000276:immunology* / Q000494:pharmacology; D000916:Antibody Diversity; D000937:Antigen-Antibody Reactions; D001483:Base Sequence; D001963:Bromelains; D002460:Cell Line; D004912:Erythrocytes / Q000276:immunology*; D005260:Female; D005803:Genes, Immunoglobulin; D006461:Hemolysis; D006825:Hybridomas / Q000032:analysis; D007143:Immunoglobulin Heavy Chains / Q000235:genetics; D007147:Immunoglobulin Light Chains / Q000235:genetics; D007135:Immunoglobulin Variable Region / Q000235:genetics; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D008808:Mice, Inbred CBA; D008969:Molecular Sequence Data; D000644:Quaternary Ammonium Compounds
D000911:Antibodies, Monoclonal; D007143:Immunoglobulin Heavy Chains; D007147:Immunoglobulin Light Chains; D007135:Immunoglobulin Variable Region; D000644:Quaternary Ammonium Compounds; D001963:Bromelains