Tracking the repertoire of human adult and neonatal <scp>T</scp> cells during <i>ex vivo</i> amplification

Neller, Michelle A.; Sewell, Andrew K.; Burrows, Scott R.; Miles, John J.

doi:10.1111/bjh.12022

Cited by 4 publications

(5 citation statements)

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“…Lymphocytes were isolated by Ficoll‐Paque PLUS (GE Healthcare, Little Chalfont, UK) density gradient centrifugation and cryopreserved in R10 medium (RPMI‐1640 containing 10% FBS) supplemented with 10% DMSO (Sigma‐Aldrich, Gillingham, UK). For some UCB samples, cells were expanded by adding Human T‐Activator CD3/CD28 Dynabeads (Life Technologies, Paisley, UK) as described previously 40 . All samples were obtained with written consent and all protocols were approved by ethics committees at The Anthony Nolan Trust and the Queensland Institute of Medical Research.…”

Section: Methodsmentioning

confidence: 99%

“…For some UCB samples, cells were expanded by adding Human T-Activator CD3/CD28 Dynabeads (Life Technologies, Paisley, UK) as described previously. 40 All samples were obtained with written consent and all protocols were approved by ethics committees at The Anthony Nolan Trust and the Queensland Institute of Medical Research.…”

Section: Blood Samples and Processingmentioning

confidence: 99%

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Naive CD8⁺ T‐cell precursors display structured TCR repertoires and composite antigen‐driven selection dynamics

et al. 2015

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Basic parameters of the naive antigen (Ag)-specific T-cell repertoire in humans remain poorly defined. Systematic characterization of this ‘ground state' immunity in comparison with memory will allow a better understanding of clonal selection during immune challenge. Here, we used high-definition cell isolation from umbilical cord blood samples to establish the baseline frequency, phenotype and T-cell antigen receptor (TCR) repertoire of CD8+ T-cell precursor populations specific for a range of viral and self-derived Ags. Across the board, these precursor populations were phenotypically naive and occurred with hierarchical frequencies clustered by Ag specificity. The corresponding patterns of TCR architecture were highly ordered and displayed partial overlap with adult memory, indicating biased structuring of the T-cell repertoire during Ag-driven selection. Collectively, these results provide new insights into the complex nature and dynamics of the naive T-cell compartment.

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Section: Methodsmentioning

confidence: 99%

Section: Blood Samples and Processingmentioning

confidence: 99%

Naive CD8⁺ T‐cell precursors display structured TCR repertoires and composite antigen‐driven selection dynamics

et al. 2015

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“…Previous studies have applied a T-cell library approach to study T-cell frequencies, but instead used PHA in combination with irradiated allogeneic feeder cells for T-cell expansion ( Campion et al, 2014 , Geiger et al, 2009 ). The CD3/CD28 beads used in our strategy have been shown to better preserve the TCR repertoire during in vitro expansion ( Neller et al, 2012 ). Nevertheless, while this methodology maintains the general TRBV families and dominant antigen-specific T-cell responses faithfully, it remains possible that extremely rare clones are lost during this expansion phase.…”

Section: Discussionmentioning

confidence: 99%

T-cell libraries allow simple parallel generation of multiple peptide-specific human T-cell clones

Theaker

Rius

Greenshields‐Watson

et al. 2016

Journal of Immunological Methods

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Isolation of peptide-specific T-cell clones is highly desirable for determining the role of T-cells in human disease, as well as for the development of therapies and diagnostics. However, generation of monoclonal T-cells with the required specificity is challenging and time-consuming. Here we describe a library-based strategy for the simple parallel detection and isolation of multiple peptide-specific human T-cell clones from CD8+ or CD4+ polyclonal T-cell populations. T-cells were first amplified by CD3/CD28 microbeads in a 96U-well library format, prior to screening for desired peptide recognition. T-cells from peptide-reactive wells were then subjected to cytokine-mediated enrichment followed by single-cell cloning, with the entire process from sample to validated clone taking as little as 6 weeks. Overall, T-cell libraries represent an efficient and relatively rapid tool for the generation of peptide-specific T-cell clones, with applications shown here in infectious disease (Epstein–Barr virus, influenza A, and Ebola virus), autoimmunity (type 1 diabetes) and cancer.

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“…Expansion to generate large enough input can alter population profiles. [9][10][11] We recently reported on the use of magnetic iron-oxide nanoparticles (NPs) to support the presentation of >10 4 pMHC tetramers per particle (Peng et al, under revision). These high avidity pMHCpresenting NPs (pNPs), which can also be labeled with multi-functional DNA barcodes, permit an improved efficiency of antigen-specific T cell capture relative to pMHC tetramers.…”

Section: Introductionmentioning

confidence: 99%

MATE-Seq: microfluidic antigen-TCR engagement sequencing

Peng

et al. 2019

Lab Chip

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Adaptive immunity is based on peptide antigen recognition. Our ability to harness the immune system for therapeutic gain relies on the discovery of the T cell receptor (TCR) genes that selectively target antigens from infections, mutated proteins, and foreign agents. Here we present a method that selectively labels peptide antigen-specific CD8+ T-cells in human blood using magnetic nanoparticles functionalized with peptide-MHC tetramers, isolates these specific cells within an integrated microfluidic device, and directly amplifies the TCR genes for sequencing. Critically, the identity of the peptide recognized by the TCR is preserved, providing the link between peptide and gene. The platform requires inputs on the order of just 100,000 CD8+ T cells, can be multiplexed for simultaneous analysis of multiple peptides, and performs sorting and isolation on chip. We demonstrate 1000-fold sensitivity enhancement of antigen-specific Tcell receptor detection and simultaneous capture of two virus antigen-specific T-cell receptors from samples of human blood.

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Tracking the repertoire of human adult and neonatal T cells during ex vivo amplification

Cited by 4 publications

References 5 publications

Naive CD8⁺ T‐cell precursors display structured TCR repertoires and composite antigen‐driven selection dynamics

Naive CD8⁺ T‐cell precursors display structured TCR repertoires and composite antigen‐driven selection dynamics

T-cell libraries allow simple parallel generation of multiple peptide-specific human T-cell clones

MATE-Seq: microfluidic antigen-TCR engagement sequencing

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Tracking the repertoire of human adult and neonatal T cells during ex vivo amplification

Cited by 4 publications

References 5 publications

Naive CD8+ T‐cell precursors display structured TCR repertoires and composite antigen‐driven selection dynamics

Naive CD8+ T‐cell precursors display structured TCR repertoires and composite antigen‐driven selection dynamics

T-cell libraries allow simple parallel generation of multiple peptide-specific human T-cell clones

MATE-Seq: microfluidic antigen-TCR engagement sequencing

Contact Info

Product

Resources

About

Naive CD8⁺ T‐cell precursors display structured TCR repertoires and composite antigen‐driven selection dynamics

Naive CD8⁺ T‐cell precursors display structured TCR repertoires and composite antigen‐driven selection dynamics