Detection of one VH antibody sequence in both healthy donors and urticaria patients.
We have previously isolated anti-FcepsilonRIalpha autoantibodies from phage libraries of healthy donors and urticaria patients. Strikingly, the same antibody, LTMalpha15, was isolated from both libraries. Sequence analysis revealed a germline configuration of the LTMalpha15 variable heavy (V(H)) chain with a slightly mutated variable light (V(L)) chain supporting its classification as a natural autoantibody. Distribution analysis of anti-FcepsilonRIalpha autoantibodies by functional or serological tests delivered conflicting data. For this reason we have developed a new real-time PCR to analyse the distribution of LTMalpha15V(H) in healthy donors and urticaria patients. Our new bioinformatic program permitted the design of a minor groove binder (MGB) TaqMan probe that specifically detected the LTMalpha15V(H). We were able to demonstrate a broad range of rearranged V(H) gene copy number without any correlation to the state of health. Monitoring LTMalpha15V(H) gene copy number in a single donor over a period of 70 days revealed a time-related fluctuation of circulating B cells carrying LTMalpha15V(H). We propose that our real-time PCR may serve as a model for the quantification of natural antibody sequences at a monoclonal level.
Fux
Michaela
M
Institute of Immunology, University of Bern, Inselspital, Switzerland.
Vogel
Monique
M
Stadler
Michael B
MB
Stadler
Beda M
BM
Miescher
Sylvia M
SM
D001323:Autoantibodies / Q000235:genetics* / Q000276:immunology; D001402:B-Lymphocytes / Q000276:immunology / Q000378:metabolism; D001483:Base Sequence; D022801:Complementarity Determining Regions / Q000235:genetics; D019295:Computational Biology / Q000379:methods; D017931:DNA Primers / Q000235:genetics; D015342:DNA Probes / Q000235:genetics; D018628:Gene Dosage; D015326:Gene Rearrangement, B-Lymphocyte, Heavy Chain / Q000235:genetics; D006801:Humans; D007143:Immunoglobulin Heavy Chains / Q000235:genetics*; D007135:Immunoglobulin Variable Region / Q000235:genetics*; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000379:methods; D017455:Receptors, IgE / Q000235:genetics / Q000276:immunology; D017422:Sequence Analysis, DNA / Q000379:methods*; D013997:Time Factors; D014581:Urticaria / Q000235:genetics / Q000276:immunology*
D001323:Autoantibodies; D022801:Complementarity Determining Regions; D017931:DNA Primers; D015342:DNA Probes; C475664:FCER1A protein, human; D007143:Immunoglobulin Heavy Chains; D007135:Immunoglobulin Variable Region; D017455:Receptors, IgE