Exhaustive T-cell repertoire sequencing of human peripheral blood samples reveals signatures of antigen selection and a directly measured repertoire size of at least 1 million clonotypes
René L. Warren,J. Douglas Freeman,Thomas Zeng,Gina Choe,Sarah A. Munro,Richard D. Moore,John R. Webb,Robert A. Holt +7 more
TLDR
It is shown that the sensitivity of sequence-based repertoire profiling is limited by both sequencing depth and sequencing accuracy, and a new, directly measured, lower limit on individual T-cell repertoire size is established.Abstract:
Massively parallel sequencing is a useful approach for characterizing T-cell receptor diversity. However, immune receptors are extraordinarily difficult sequencing targets because any given receptor variant may be present in very low abundance and may differ legitimately by only a single nucleotide. We show that the sensitivity of sequence-based repertoire profiling is limited by both sequencing depth and sequencing accuracy. At two timepoints, 1 wk apart, we isolated bulk PBMC plus naive (CD45RA+/CD45RO-) and memory (CD45RA-/CD45RO+) T-cell subsets from a healthy donor. From T-cell receptor beta chain (TCRB) mRNA we constructed and sequenced multiple libraries to obtain a total of 1.7 billion paired sequence reads. The sequencing error rate was determined empirically and used to inform a high stringency data filtering procedure. The error filtered data yielded 1,061,522 distinct TCRB nucleotide sequences from this subject which establishes a new, directly measured, lower limit on individual T-cell repertoire size and provides a useful reference set of sequences for repertoire analysis. TCRB nucleotide sequences obtained from two additional donors were compared to those from the first donor and revealed limited sharing (up to 1.1%) of nucleotide sequences among donors, but substantially higher sharing (up to 14.2%) of inferred amino acid sequences. For each donor, shared amino acid sequences were encoded by a much larger diversity of nucleotide sequences than were unshared amino acid sequences. We also observed a highly statistically significant association between numbers of shared sequences and shared HLA class I alleles.read more
Citations
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Journal ArticleDOI
How many TCR clonotypes does a body maintain
TL;DR: A computational model of homeostasis of a multiclonal population of T cells with self pMHC stimuli is devised and analysed, finding a constant mean total number of cells is maintained by a balance between cell division and death, and a stable number of clonotypes by a Balance between thymic production of new clonotype and extinction of existing ones.
Sequence analysis of T-cell repertoires in health
TL;DR: This nascent field and TCR-seq methodology, which involves the use of next generation sequencing platforms to generate large numbers of short DNA sequences covering key regions of the TCR coding sequence, is introduced and discusses recent insights into healthy and diseased TCR repertoires.
Journal ArticleDOI
TCR repertoire analysis by next generation sequencing allows complex differential diagnosis of T cell-related pathology.
Mikalai Dziubianau,Jochen Hecht,Leon Kuchenbecker,Arne Sattler,Ulrik Stervbo,Christian Rödelsperger,Peter Nickel,Avidan U. Neumann,Peter N. Robinson,Stefan Mundlos,H-D Volk,Andreas Thiel,Petra Reinke,Nina Babel +13 more
TL;DR: A next generation sequencing (NGS)‐based platform for the highly quantitative clonotype characterization of T cells is developed and the established approach will provide insights into the regulation of virus‐specific/anti‐tumor immunity and has high diagnostic potential in the clinical routine.
Journal ArticleDOI
Best practices for bioinformatic characterization of neoantigens for clinical utility
Megan Richters,Huiming Xia,Katie M. Campbell,William E. Gillanders,Obi L. Griffith,Malachi Griffith +5 more
TL;DR: This article provides practical guidance, specific recommendations, and extensive discussion of critical concepts and points of confusion in the practice of neoantigen characterization for clinical use, and outlines necessary areas of development.
Journal ArticleDOI
Dynamics of Individual T Cell Repertoires: From Cord Blood to Centenarians
Olga V. Britanova,Mikhail Shugay,Ekaterina M. Merzlyak,Dmitriy B. Staroverov,Ekaterina V. Putintseva,Maria A. Turchaninova,Ilgar Z. Mamedov,Mikhail V. Pogorelyy,Dmitriy A. Bolotin,Mark Izraelson,Alexey N. Davydov,Evgeny S. Egorov,Sofya A. Kasatskaya,Denis V. Rebrikov,Sergey A. Lukyanov,Dmitriy M. Chudakov +15 more
TL;DR: It is shown that the fetal TCR repertoire, albeit structurally maintained within regulated borders due to the lower numbers of randomly added nucleotides, is not limited with respect to observed functional diversity, and a more general explanation for the previous observations on the relationships between longevity and immunity is proposed.
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