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

Brandon J. Dekosky; Gregory C. Ippolito; Ryan P. Deschner; Jason J. Lavinder; Yariv Wine; Brandon M. Rawlings; Navin Varadarajan; Claudia Giesecke; Thomas Dörner; Sarah F. Andrews; Patrick C. Wilson; Scott P. Hunicke-Smith; C. Grant Willson; Andrew D. Ellington; George Georgiou

doi:10.1038/nbt.2492

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

Brandon J. Dekosky, Gregory C. Ippolito, Ryan P. Deschner, Jason J. Lavinder, Yariv Wine, Brandon M. Rawlings, Navin Varadarajan, Claudia Giesecke, Thomas Dörner, Sarah F. Andrews, Patrick C. Wilson, Scott P. Hunicke-Smith, C. Grant Willson, Andrew D. Ellington, George Georgiou

Research output: Contribution to journal › Article › peer-review

Abstract

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 × 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.

Original language	English
Pages (from-to)	166-169
Number of pages	4
Journal	Nature biotechnology
Volume	31
Issue number	2
DOIs	https://doi.org/10.1038/nbt.2492
State	Published - Feb 2013
Externally published	Yes

All Science Journal Classification (ASJC) codes

Biotechnology
Bioengineering
Applied Microbiology and Biotechnology
Molecular Medicine
Biomedical Engineering

UN SDGs

This output contributes to the following UN Sustainable Development Goals (SDGs)

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10.1038/nbt.2492

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Dekosky, B. J., Ippolito, G. C., Deschner, R. P., Lavinder, J. J., Wine, Y., Rawlings, B. M., Varadarajan, N., Giesecke, C., Dörner, T., Andrews, S. F., Wilson, P. C., Hunicke-Smith, S. P., Willson, C. G., Ellington, A. D., & Georgiou, G. (2013). High-throughput sequencing of the paired human immunoglobulin heavy and light chain repertoire. Nature biotechnology, 31(2), 166-169. https://doi.org/10.1038/nbt.2492

@article{78a4970165fe4f5b85a0b85a284233ad,

title = "High-throughput sequencing of the paired human immunoglobulin heavy and light chain repertoire",

abstract = "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 × 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.",

author = "Dekosky, \{Brandon J.\} and Ippolito, \{Gregory C.\} and Deschner, \{Ryan P.\} and Lavinder, \{Jason J.\} and Yariv Wine and Rawlings, \{Brandon M.\} and Navin Varadarajan and Claudia Giesecke and Thomas D{\"o}rner and Andrews, \{Sarah F.\} and Wilson, \{Patrick C.\} and Hunicke-Smith, \{Scott P.\} and Willson, \{C. Grant\} and Ellington, \{Andrew D.\} and George Georgiou",

note = "Funding Information: We thank B. Iverson and M. Pogson for insightful discussions and thoughtful advice, C. Das for assistance with antibody expression, and S. Reddy for help with initial studies. This work was funded by fellowships to B.J.D. from the Hertz Foundation, the University of Texas Donald D. Harrington Foundation and the National Science Foundation, and also by the US National Institutes of Health U19AI057234-09 (P.C.W.), U19AI057234-09 (G.G.), and U54AI057156 (G.G. and A.E.D.).",

year = "2013",

month = feb,

doi = "10.1038/nbt.2492",

language = "الإنجليزيّة",

volume = "31",

pages = "166--169",

journal = "Nature biotechnology",

issn = "1087-0156",

publisher = "Nature Research",

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Dekosky, BJ, Ippolito, GC, Deschner, RP, Lavinder, JJ, Wine, Y, Rawlings, BM, Varadarajan, N, Giesecke, C, Dörner, T, Andrews, SF, Wilson, PC, Hunicke-Smith, SP, Willson, CG, Ellington, AD & Georgiou, G 2013, 'High-throughput sequencing of the paired human immunoglobulin heavy and light chain repertoire', Nature biotechnology, vol. 31, no. 2, pp. 166-169. https://doi.org/10.1038/nbt.2492

TY - JOUR

T1 - High-throughput sequencing of the paired human immunoglobulin heavy and light chain repertoire

AU - Dekosky, Brandon J.

AU - Ippolito, Gregory C.

AU - Deschner, Ryan P.

AU - Lavinder, Jason J.

AU - Wine, Yariv

AU - Rawlings, Brandon M.

AU - Varadarajan, Navin

AU - Giesecke, Claudia

AU - Dörner, Thomas

AU - Andrews, Sarah F.

AU - Wilson, Patrick C.

AU - Hunicke-Smith, Scott P.

AU - Willson, C. Grant

AU - Ellington, Andrew D.

AU - Georgiou, George

N1 - Funding Information: We thank B. Iverson and M. Pogson for insightful discussions and thoughtful advice, C. Das for assistance with antibody expression, and S. Reddy for help with initial studies. This work was funded by fellowships to B.J.D. from the Hertz Foundation, the University of Texas Donald D. Harrington Foundation and the National Science Foundation, and also by the US National Institutes of Health U19AI057234-09 (P.C.W.), U19AI057234-09 (G.G.), and U54AI057156 (G.G. and A.E.D.).

PY - 2013/2

Y1 - 2013/2

N2 - 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 × 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.

AB - 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 × 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.

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U2 - 10.1038/nbt.2492

DO - 10.1038/nbt.2492

M3 - مقالة

SN - 1087-0156

VL - 31

SP - 166

EP - 169

JO - Nature biotechnology

JF - Nature biotechnology

IS - 2

ER -

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

Abstract

All Science Journal Classification (ASJC) codes

UN SDGs

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