![]() The Generation of Antibody Diversity (Garland Science, New York, 2002).īoyd, S. Clonal selection and learning in the antibody system. This dataset enables genetic study of the baseline human antibody repertoire at an unprecedented depth and granularity, which reveals largely unique repertoires for each individual studied, a subpopulation of universally shared antibody clonotypes, and an exceptional overall diversity of the antibody repertoire. Here we examine the circulating B cell populations of ten human subjects and present what is, to our knowledge, the largest single collection of adaptive immune receptor sequences described to date, comprising almost 3 billion antibody heavy-chain sequences. Furthermore, because much of the B lymphocyte population is localized in organs or tissues that cannot be comprehensively sampled from living subjects, human repertoire studies have focused on circulating B cells 3. The amount of information encoded by all of the rearranged antibody and T cell receptor genes in one person-the ‘genome’ of the adaptive immune system-exceeds the size of the human genome by more than four orders of magnitude. Full-scale analyses of human antibody repertoires have been prohibitively difficult, primarily owing to their massive size. Because the number of peripheral blood B cells in a healthy adult human is on the order of 5 × 10 9, the circulating B cell population samples only a small fraction of this diversity. The diversity of the naive antibody repertoire in humans is estimated to be at least 10 12 unique antibodies 2. This flexibility is achieved by the presence of a large repertoire of naive antibodies, the diversity of which is expanded by somatic hypermutation following antigen exposure 1. These specific antibodies should facilitate further characterization of the functionally pleiotropic viral rep proteins.In principle, humans can produce an antibody response to any non-self-antigen molecule in the appropriate context. coli are capable of detecting wild-type AAV rep proteins in virus-infected mammalian cells. These results demonstrate that antisera raised against an AAV rep protein synthesized in E. Immunofluorescence analysis of AAV-infected human cells revealed that the rep proteins are localized primarily in the nucleus of the infected cell and have a distribution different from that of AAV capsid protein. The antibodies also recognized novel forms of the rep proteins expressed from mutant AAV genomes. These new rep proteins originate from the transcription promoter at map unit 19 in the AAV genome and may indicate use of alternate AUG codons or protein modification. These antibodies were capable of detecting all four AAV rep proteins in human cells transfected with AAV-containing plasmids as well as new species of 47 and 35 kDa in molecular weight. coli, rep 78.93, was used to raise specific antibodies in rabbits. We have constructed a prokaryotic vector which expressed in Escherichia coli a region of AAV comprising 93% of the largest AAV rep protein. At least four overlapping polypeptides are expressed from the rep gene. The rep gene of the defective human parvovirus, adeno-associated virus, (AAV) mediates several trans-acting functions important to virus replication, transcription, and gene expression. ![]()
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