Quality of antibodies secreted by clones in microcultures from B cells enriched on haptenated gelatin: isotypes and avidities.
Populations of murine B cells enriched for fluorescein (FLU)- or phosphocholine (PC)-binding cells stimulated with LPS, or FLU- or PC-LPS at low density in 10 microliter cultures form clones of cells that secrete antibodies. Antibody isotypes were determined by radioimmunoassay and their avidities were determined relative to standard, monoclonal antibodies by hapten inhibition using a radioimmunoassay. These analyses further characterize the development of B cell clones in microcultures and reveal that differing culturing conditions stimulate qualitatively different B cell populations to divide and differentiate. Without filler cells, isotype switching is rare. Co-culturing B cells with 10(5) (CBA/N x BALB/c) F1 male thymocyte filler cells leads to IgG and/or IgA antibody secretion by 15-20% of cultures; antibodies from clones that switch isotypes are exclusively of high avidity. IgM is almost always present as one clonal product; pre-switched cells rarely score in microcultures. Without filler cells, a high percentage of antibodies from FLU-LPS stimulated, FLU-binding cells are of high avidity (60%). However, clonotypes of lower avidity dominate with mitogenic culture conditions, 100 micrograms/ml LPS or with thymocytes. PC-binding cells are less sensitive to these mitogenic effects. Antibodies produced by PC-specific clones have a more restricted pattern of avidities and resemble in quality anti-PC antibodies produced in vivo.
Schweitzer
P A
PA
Department of Biology, University of Pennsylvania, Philadelphia 19104-6018.
Cebra
J J
JJ
D000818:Animals; D000915:Antibody Affinity; D001402:B-Lymphocytes / Q000276:immunology*; D002478:Cells, Cultured; D002999:Clone Cells / Q000276:immunology; D005780:Gelatin; D006241:Haptens / Q000276:immunology*; D007132:Immunoglobulin Isotypes / Q000032:analysis*; D007136:Immunoglobulins / Q000032:analysis / Q000096:biosynthesis*; D008297:Male; D051379:Mice; D008807:Mice, Inbred BALB C; D008808:Mice, Inbred CBA
D006241:Haptens; D007132:Immunoglobulin Isotypes; D007136:Immunoglobulins; D005780:Gelatin