Precise determination of the diversity of a combinatorial antibody library gives insight into the human immunoglobulin repertoire
Jacob Glanville,Wenwu Zhai,Jan Berka,Dilduz Telman,Gabriella Huerta,Gautam R. Mehta,Irene Ni,Li Mei,Purnima Sundar,Giles Day,David Cox,Arvind Rajpal,Jaume Pons +12 more
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TLDR
A general method for assessing human antibody sequence diversity displayed on phage using massively parallel pyrosequencing, a novel application of Kabat column-labeled profile Hidden Markov Models, and translated complementarity determining region (CDR) capture-recapture analysis is presented.Abstract:
Antibody repertoire diversity, potentially as high as 1011 unique molecules in a single individual, confounds characterization by conventional sequence analyses. In this study, we present a general method for assessing human antibody sequence diversity displayed on phage using massively parallel pyrosequencing, a novel application of Kabat column-labeled profile Hidden Markov Models, and translated complementarity determining region (CDR) capture-recapture analysis. Pyrosequencing of domain amplicon and RCA PCR products generated 1.5 × 106 reads, including more than 1.9 × 105 high quality, full-length sequences of antibody variable fragment (Fv) variable domains. Novel methods for germline and CDR classification and fine characterization of sequence diversity in the 6 CDRs are presented. Diverse germline contributions to the repertoire with random heavy and light chain pairing are observed. All germline families were found to be represented in 1.7 × 104 sequences obtained from repeated panning of the library. While the most variable CDR (CDR-H3) presents significant length and sequence variability, we find a substantial contribution to total diversity from somatically mutated germline encoded CDRs 1 and 2. Using a capture-recapture method, the total diversity of the antibody library obtained from a human donor Immunoglobulin M (IgM) pool was determined to be at least 3.5 × 1010. The results provide insights into the role of IgM diversification, display library construction, and productive germline usages in antibody libraries and the humoral repertoire.read more
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Focused evolution of hiv-1 neutralizing antibodies revealed by crystal structures and deep sequencing
TL;DR: In this article, the authors used X-ray crystallography and 454 pyrosequencing to characterize additional antibodies from HIV-1-infected individuals, revealing a convergent mode of binding of different antibodies to the same CD4-binding-site epitope.
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TL;DR: The GLIPH algorithm can analyse large numbers of TCR sequences and define TCR specificity groups shared by TCRs and individuals, which should greatly accelerate the analysis of T cell responses and expedite the identification of specific ligands.
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Developmental pathway for potent V1V2-directed HIV-neutralizing antibodies.
Nicole A. Doria-Rose,Chaim A. Schramm,Jason Gorman,Penny L. Moore,Jinal N. Bhiman,Brandon J. DeKosky,Michael J. Ernandes,Ivelin S. Georgiev,Helen J. Kim,Marie Pancera,Ryan P. Staupe,Han Altae-Tran,Robert T. Bailer,Ema T. Crooks,Albert Cupo,Aliaksandr Druz,Nigel Garrett,Kam Hon Hoi,Rui Kong,Mark K. Louder,Nancy S. Longo,Krisha McKee,Molati Nonyane,Sijy O'Dell,Ryan S. Roark,Rebecca S. Rudicell,Stephen D. Schmidt,Daniel J. Sheward,Cinque Soto,Constantinos Kurt Wibmer,Yongping Yang,Zhenhai Zhang,Nisc Comparative Sequencing,James C. Mullikin,James M. Binley,Rogier W. Sanders,Ian A. Wilson,John P. Moore,Andrew B. Ward,George Georgiou,Carolyn Williamson,Salim S. Abdool Karim,Lynn Morris,Peter D. Kwong,Lawrence Shapiro,John R. Mascola +45 more
TL;DR: HIV-1 V1V2-directed neutralizing antibodies can develop relatively rapidly through initial selection of B cells with a long CDR H3, and limited subsequent somatic hypermutation, and provide important insights relevant to HIV-1 vaccine development.
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George Georgiou,Gregory C. Ippolito,John F. Beausang,Christian E. Busse,Hedda Wardemann,Stephen R. Quake +5 more
TL;DR: In this paper, a standardized experimental design framework that will enable the sharing and meta-analysis of sequencing data generated by different laboratories is proposed, which can be applied to detect B-cell malignancies with high sensitivity, to discover antibodies specific for antigens of interest, to guide vaccine development and to understand autoimmunity.
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Immunoglobulin analysis tool: a novel tool for the analysis of human and mouse heavy and light chain transcripts.
Tobias Rogosch,Sebastian Kerzel,Kam Hon Hoi,Zhixin Zhang,Rolf F. Maier,Gregory C. Ippolito,Michael Zemlin +6 more
TL;DR: IgAT is a useful tool to gain insights into the selective forces and functional properties of small to extremely large collections of Ig transcripts, thereby assisting a researcher to mine a data set to its fullest.
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TL;DR: A novel bacteriophage lambda vector system was used to express in Escherichia coli a combinatorial library of Fab fragments of the mouse antibody repertoire, which allows rapid and easy identification of monoclonal Fab fragments in a form suitable for genetic manipulation.
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