title: Frequency analysis of functional immunoglobulin C- and V-gene expression by mitogen-reactive B cells in germfree mice fed chemically defined ultra-filtered "antigen-free" diet.
abstract: The frequencies of lipopolysaccharide (LPS)-reactive B cells and their antibody specificity repertoire have been determined in the spleen and bone marrow (BM) of conventional (CV) and "antigen-free" C3H/HeCr mice of various ages. The antigen-free mice were germfree (GF)-raised and were fed an ultrafiltered solution of chemically defined (CD) low m.w. nutrients, and were thus devoid of exogenous antigenic stimulation. Spleen and BM cells were grown in a limiting dilution culture system that allows the growth and development of every newly formed LPS-reactive B cell into a clone of IgM-secreting cells which are capable of switching to other immunoglobulin (Ig) heavy chain isotypes (C-gene expression). The secretion of IgM and IgG1 was determined in the protein A plaque assay, whereas specific IgM antibody-secreting cells (V-gene expression) were detected in plaque assays specific for various heterologous erythrocytes and sheep red blood cells (SRBC) coupled with a number of different haptens. The absolute frequency of LPS-reactive B cells and their capacity to switch to IgG1-secretion was not significantly different in 8- to 12-wk-old and 52-wk-old GF-CD mice and their age-matched CV controls. Moreover, no differences were observed in the frequencies of antigen-specific B cells within the pool of LPS reactive B cells. These frequencies ranged from 1 in 20 to 1 in 50 for NIP4-SRBC and NNP2-SRBC, from 1 in 100 to 1 in 150 for NIP0.4-SRBC, from 1 in 50 to 1 in 100 for TNP30-SRBC, and from 1 in 1000 to 1 in 2000 for SRBC and horse red blood cells. Within the limitations of having determined the switching capacity of IgM to IgG1 only and having assessed only a minor fraction of the total B cell antibody-specificity repertoire, the data indicate that young and old GF-CD mice, although devoid of exogenous antigenic and/or mitogenic stimulation, generate B cells with a similar switching capacity and a similar IgM antibody specificity repertoire as CV mice.
mesh_terms: D000818:Animals; D001402:B-Lymphocytes / Q000276:immunology* / Q000378:metabolism / Q000502:physiology; D001854:Bone Marrow Cells; D002452:Cell Count; D005260:Female; D005526:Food, Formulated; D005615:Freezing; D005856:Germ-Free Life; D007127:Immunoglobulin Constant Regions / Q000235:genetics*; D007075:Immunoglobulin M / Q000096:biosynthesis; D007135:Immunoglobulin Variable Region / Q000235:genetics*; D007136:Immunoglobulins / Q000235:genetics*; D008070:Lipopolysaccharides / Q000494:pharmacology; D008213:Lymphocyte Activation; D008297:Male; D051379:Mice; D008809:Mice, Inbred C3H; D008810:Mice, Inbred C57BL; D051381:Rats; D011917:Rats, Inbred Lew; D013154:Spleen / Q000166:cytology
chemical_list: D007127:Immunoglobulin Constant Regions; D007075:Immunoglobulin M; D007135:Immunoglobulin Variable Region; D007136:Immunoglobulins; D008070:Lipopolysaccharides