High-throughput sequencing of the paired human immunoglobulin heavy and light chain repertoire

被引：328

作者：

DeKosky, Brandon J. ^{[1
]}

Ippolito, Gregory C. ^{[2
]}

Deschner, Ryan P. ^{[1
]}

Lavinder, Jason J. ^{[3
]}

Wine, Yariv ^{[1
]}

Rawlings, Brandon M. ^{[1
]}

Varadarajan, Navin ^{[4
]}

Giesecke, Claudia ^{[5
,6
]}

Doerner, Thomas ^{[5
,6
]}

Andrews, Sarah F. ^{[7
]}

Wilson, Patrick C. ^{[7
]}

Hunicke-Smith, Scott P. ^{[3
]}

Willson, C. Grant ^{[1
,8
]}

Ellington, Andrew D. ^{[3
,8
]}

Georgiou, George ^{[1
,2
,3
,9
]}

机构：

[1] Univ Texas Austin, Dept Chem Engn, Austin, TX 78712 USA

[2] Univ Texas Austin, Sect Mol Genet & Microbiol, Austin, TX 78712 USA

[3] Univ Texas Austin, Inst Cellular & Mol Biol, Austin, TX 78712 USA

[4] Univ Houston, Dept Chem & Biomol Engn, Houston, TX USA

[5] Charite, Dept Med Rheumatol & Clin Immunol, D-13353 Berlin, Germany

[6] German Rheumatism Res Ctr DRFZ, Berlin, Germany

[7] Univ Chicago, Dept Med, Rheumatol Sect, Chicago, IL 60637 USA

[8] Univ Texas Austin, Dept Chem & Biochem, Austin, TX 78712 USA

[9] Univ Texas Austin, Dept Biomed Engn, Austin, TX 78712 USA

来源：

NATURE BIOTECHNOLOGY | 2013年 / 31卷 / 02期

基金：

美国国家科学基金会; 美国国家卫生研究院;

关键词：

HUMAN MONOCLONAL-ANTIBODIES; MEMORY B-CELLS; PLASMA-CELLS; NEXT-GENERATION; CLONING; ANTIGEN;

D O I：

10.1038/nbt.2492

中图分类号：

Q81 [生物工程学（生物技术）]; Q93 [微生物学];

学科分类号：

071005 ; 0836 ; 090102 ; 100705 ;

摘要：

Each B-cell receptor consists of a pair of heavy and light chains. High-throughput sequencing can identify large numbers of heavy- and light-chain variable regions (V-H and V-L) in a given B-cell repertoire, but information about endogenous pairing of heavy and light chains is lost after bulk lysis of B-cell populations. Here we describe a way to retain this pairing information. In our approach, single B cells (>5 x 10(4) capacity per experiment) are deposited in a high-density microwell plate (125 pl/well) and lysed in situ. mRNA is then captured on magnetic beads, reverse transcribed and amplified by emulsion V-H:V-L linkage PCR. The linked transcripts are analyzed by Illumina high-throughput sequencing. We validated the fidelity of V-H:V-L pairs identified by this approach and used the method to sequence the repertoire of three human cell subsets-peripheral blood IgG(+) B cells, peripheral plasmablasts isolated after tetanus toxoid immunization and memory B cells isolated after seasonal influenza vaccination.

引用

页码：166 / 169

页数：4

共 23 条

[1] Rep-Seq: uncovering the immunological repertoire through next-generation sequencing [J].

Benichou, Jennifer ;

Ben-Hamo, Rotem ;

Louzoun, Yoram ;

Efroni, Sol .

IMMUNOLOGY, 2012, 135 (03) :183-191

[2] A proteomics approach for the identification and cloning of monoclonal antibodies from serum [J].

Cheung, Wan Cheung ;

Beausoleil, Sean A. ;

Zhang, Xiaowu ;

Sato, Shuji ;

Schieferl, Sandra M. ;

Wieler, James S. ;

Beaudet, Jason G. ;

Ramenani, Ravi K. ;

Popova, Lana ;

Comb, Michael J. ;

Rush, John ;

Polakiewicz, Roberto D. .

NATURE BIOTECHNOLOGY, 2012, 30 (05) :447-+

[3] Sequencing antibody repertoires The next generation [J].

Fischer, Nicolas .

MABS, 2011, 3 (01) :17-20

[4] Secondary Immunization Generates Clonally Related Antigen-Specific Plasma Cells and Memory B Cells [J].

Froelich, Daniela ;

Giesecke, Claudia ;

Mei, Henrik E. ;

Reiter, Karin ;

Daridon, Capucine ;

Lipsky, Peter E. ;

Doerner, Thomas .

JOURNAL OF IMMUNOLOGY, 2010, 185 (05) :3103-3110

[5] Naive antibody gene-segment frequencies are heritable and unaltered by chronic lymphocyte ablation [J].

Glanville, Jacob ;

Kuo, Tracy C. ;

von Buedingen, H. -Christian ;

Guey, Lin ;

Berka, Jan ;

Sundar, Purnima D. ;

Huerta, Gabriella ;

Mehta, Gautam R. ;

Oksenberg, Jorge R. ;

Hauser, Stephen L. ;

Cox, David R. ;

Rajpal, Arvind ;

Pons, Jaume .

PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 2011, 108 (50) :20066-20071

[6] Antibody Repertoires in Humanized NOD-scid-IL2Rγnull Mice and Human B Cells Reveals Human-Like Diversification and Tolerance Checkpoints in the Mouse [J].

Ippolito, Gregory C. ;

Hoi, Kam Hon ;

Reddy, Sai T. ;

Carroll, Sean M. ;

Ge, Xin ;

Rogosch, Tobias ;

Zemlin, Michael ;

Shultz, Leonard D. ;

Ellington, Andrew D. ;

VanDenBerg, Carla L. ;

Georgiou, George .

PLOS ONE, 2012, 7 (04)

[7] Pandemic H1N1 influenza vaccine induces a recall response in humans that favors broadly cross-reactive memory B cells [J].

Li, Gui-Mei ;

Chiu, Christopher ;

Wrammert, Jens ;

McCausland, Megan ;

Andrews, Sarah F. ;

Zheng, Nai-Ying ;

Lee, Jane-Hwei ;

Huang, Min ;

Qu, Xinyan ;

Edupuganti, Srilatha ;

Mulligan, Mark ;

Das, Suman R. ;

Yewdell, Jonathan W. ;

Mehta, Aneesh K. ;

Wilson, Patrick C. ;

Ahmed, Rafi .

PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 2012, 109 (23) :9047-9052

[8] Isolation of human antibody repertoires with preservation of the natural heavy and light chain pairing [J].

Meijer, PJ ;

Andersen, PS ;

Hansen, MH ;

Steinaa, L ;

Jensen, A ;

Lantto, J ;

Oleksiewicz, MB ;

Tengbjerg, K ;

Poulsen, TR ;

Coljee, VW ;

Bregenholt, S ;

Haurum, JS ;

Nielsen, LS .

JOURNAL OF MOLECULAR BIOLOGY, 2006, 358 (03) :764-772

[9] Systems analysis of adaptive immunity by utilization of high-throughput technologies [J].

Reddy, Sai T. ;

Georgiou, George .

CURRENT OPINION IN BIOTECHNOLOGY, 2011, 22 (04) :584-589

[10] Monoclonal antibodies isolated without screening by analyzing the variable-gene repertoire of plasma cells [J].

Reddy, Sai T. ;

Ge, Xin ;

Miklos, Aleksandr E. ;

Hughes, Randall A. ;

Kang, Seung Hyun ;

Hoi, Kam Hon ;

Chrysostomou, Constantine ;

Hunicke-Smith, Scott P. ;

Iverson, Brent L. ;

Tucker, Philip W. ;

Ellington, Andrew D. ;

Georgiou, George .

NATURE BIOTECHNOLOGY, 2010, 28 (09) :965-U20

← 1 2 3 →