Expression Cloning of Antibodies from Single Human B Cells.

The majority of lymphomas originate from B cells at the germinal center stage. Preferential selection of B-cell clones by a limited set of antigens has been suggested to drive lymphoma development. While recent studies in B-cell chronic lymphocytic leukemia (CLL) have shown that self-reactive B-cell receptors (BCR) can generate cell-autonomous signaling and proliferation, our knowledge about the role of BCRs for the development or survival of other lymphomas remains limited.

AIRR community curation and standardised representation for immunoglobulin and T cell receptor germline sets.

Analysis of an individual's immunoglobulin or T cell receptor gene repertoire can provide important insights into immune function. High-quality analysis of adaptive immune receptor repertoire sequencing data depends upon accurate and relatively complete germline sets, but current sets are known to be incomplete. Established processes for the review and systematic naming of receptor germline genes and alleles require specific evidence and data types, but the discovery landscape is rapidly changing.

MDA-LDL vaccination induces athero-protective germinal-center-derived antibody responses.

Atherosclerosis is a chronic inflammatory disease of the arteries that can lead to thrombosis, infarction, and stroke and is the leading cause of mortality worldwide. Immunization of pro-atherogenic mice with malondialdehyde-modified low-density lipoprotein (MDA-LDL) neo-antigen is athero-protective. However, the immune response to MDA-LDL and the mechanisms responsible for this athero-protection are not completely understood.

A BALB/c IGHV Reference Set, Defined by Haplotype Analysis of Long-Read VDJ-C Sequences From F1 (BALB/c x C57BL/6) Mice.

The immunoglobulin genes of inbred mouse strains that are commonly used in models of antibody-mediated human diseases are poorly characterized. This compromises data analysis. To infer the immunoglobulin genes of BALB/c mice, we used long-read SMRT sequencing to amplify VDJ-C sequences from F1 (BALB/c x C57BL/6) hybrid animals.

Data Sharing and Reuse: A Method by the AIRR Community.

High-throughput sequencing of adaptive immune receptor repertoires (AIRR, i.e., IG and TR ) has revolutionized the ability to study the adaptive immune response via large-scale experiments.

Adaptive Immune Receptor Repertoire (AIRR) Community Guide to TR and IG Gene Annotation.

High-throughput sequencing of adaptive immune receptor repertoires (AIRR, i.e., IG and TR) has revolutionized the ability to carry out large-scale experiments to study the adaptive immune response.

An efficient single-cell based method for linking human T cell phenotype to T cell receptor sequence and specificity.

Single-cell antigen-receptor gene amplification and sequencing platforms have been used to characterize T cell receptor (TCR) repertoires but typically fail to generate paired full-length gene products for direct expression cloning and do not enable linking this data to cell phenotype information. To overcome these limitations, we established a high-throughput platform for the quantitative and qualitative analysis of human TCR repertoires that provides insights into the clonal and functional composition of human CD4 and CD8 αβ T cells at the molecular and cellular level. The strategy is a powerful tool to qualitatively assess differences between antigen receptors of phenotypically defined αβ T cell subsets, e.

ALDH4A1 is an atherosclerosis auto-antigen targeted by protective antibodies.

Cardiovascular disease (CVD) is the leading cause of mortality in the world, with most CVD-related deaths resulting from myocardial infarction or stroke. The main underlying cause of thrombosis and cardiovascular events is atherosclerosis, an inflammatory disease that can remain asymptomatic for long periods. There is an urgent need for therapeutic and diagnostic options in this area.

is an inherited risk factor for CLL through the acquisition of a single-point mutation enabling autonomous BCR signaling.

The prognosis of chronic lymphocytic leukemia (CLL) depends on different markers, including cytogenetic aberrations, oncogenic mutations, and mutational status of the immunoglobulin (Ig) heavy-chain variable (IGHV) gene. The number of IGHV mutations distinguishes mutated (M) CLL with a markedly superior prognosis from unmutated (UM) CLL cases. In addition, B cell antigen receptor (BCR) stereotypes as defined by IGHV usage and complementarity-determining regions (CDRs) classify ∼30% of CLL cases into prognostically important subsets.

A Proposed New Nomenclature for the Immunoglobulin Genes of .

Mammalian immunoglobulin (IG) genes are found in complex loci that contain hundreds of highly similar pseudogenes, functional genes and repetitive elements, which has made their investigation particularly challenging. High-throughput sequencing has provided new avenues for the investigation of these loci, and has recently been applied to study the IG genes of important inbred mouse strains, revealing unexpected differences between their IG loci. This demonstrated that the structural differences are of such magnitude that they call into question the merits of the current mouse IG gene nomenclatures.

OGRDB: a reference database of inferred immune receptor genes.

High-throughput sequencing of the adaptive immune receptor repertoire (AIRR-seq) is providing unprecedented insights into the immune response to disease and into the development of immune disorders. The accurate interpretation of AIRR-seq data depends on the existence of comprehensive germline gene reference sets. Current sets are known to be incomplete and unrepresentative of the degree of polymorphism and diversity in human and animal populations.

A comparison of immunoglobulin IGHV, IGHD and IGHJ genes in wild-derived and classical inbred mouse strains.

The genomes of classical inbred mouse strains include genes derived from all three major subspecies of the house mouse, Mus musculus. We recently posited that genetic diversity in the immunoglobulin heavy chain (IGH) gene loci of C57BL/6 and BALB/c mice reflects differences in subspecies origin. To investigate this hypothesis, we conducted high-throughput sequencing of IGH gene rearrangements to document IGH variable (IGHV), joining (IGHJ) and diversity (IGHD) genes in four inbred wild-derived mouse strains (CAST/EiJ, LEWES/EiJ, MSM/MsJ and PWD/PhJ) and a single disease model strain (NOD/ShiLtJ), collectively representing genetic backgrounds of several major mouse subspecies.

High-throughput single-cell sequencing of paired TCRα and TCRβ genes for the direct expression-cloning and functional analysis of murine T-cell receptors.

Precise clonal and functional assessments of the T cell receptor (TCR) repertoire diversity require paired TCRα and TCRβ gene sequence information at monoclonal level. However, available single-cell strategies are typically limited in throughput and often do not provide full-length DNA templates for direct gene cloning. Here, we describe a high-throughput strategy for the unbiased amplification and automated sequence analysis of paired TCRα and TCRβ genes from primary mouse T cells.

Inferred Allelic Variants of Immunoglobulin Receptor Genes: A System for Their Evaluation, Documentation, and Naming.

Immunoglobulins or antibodies are the main effector molecules of the B-cell lineage and are encoded by hundreds of variable (V), diversity (D), and joining (J) germline genes, which recombine to generate enormous IG diversity. Recently, high-throughput adaptive immune receptor repertoire sequencing (AIRR-seq) of recombined V-(D)-J genes has offered unprecedented insights into the dynamics of IG repertoires in health and disease. Faithful biological interpretation of AIRR-seq studies depends upon the annotation of raw AIRR-seq data, using reference germline gene databases to identify the germline genes within each rearrangement.

Expression Cloning of Antibodies from Single Human B Cells.

The majority of lymphomas originate from B cells at the germinal center stage. Preferential selection of B-cell clones by a limited set of antigens has been suggested to drive lymphoma development. While recent studies in chronic lymphocytic leukemia have shown that self-reactive B-cell receptors (BCR) can generate cell-autonomous signaling and proliferation, our knowledge about the role of BCRs for the development or survival of other lymphomas remains limited.

AIRR Community Standardized Representations for Annotated Immune Repertoires.

Increased interest in the immune system's involvement in pathophysiological phenomena coupled with decreased DNA sequencing costs have led to an explosion of antibody and T cell receptor sequencing data collectively termed "adaptive immune receptor repertoire sequencing" (AIRR-seq or Rep-Seq). The AIRR Community has been actively working to standardize protocols, metadata, formats, APIs, and other guidelines to promote open and reproducible studies of the immune repertoire. In this paper, we describe the work of the AIRR Community's Data Representation Working Group to develop standardized data representations for storing and sharing annotated antibody and T cell receptor data.

Clonal selection drives protective memory B cell responses in controlled human malaria infection.

Affinity maturation, the clonal selection and expansion of antigen-activated B cells expressing somatically mutated antibody variants that develop during T cell-dependent germinal center reactions, is considered pivotal for efficient development of protective B cell memory responses to infection and vaccination. Repeated antigen exposure promotes affinity maturation but each time also recruits antigen-reactive naïve B cells into the response. Here, we determined the relative impact of affinity maturation versus antigen-mediated clonal selection of naïve B cells to mount potent B cell memory responses in humans after repeated exposure to a complex pathogen, the malaria parasite (Pf).

Reproducibility and Reuse of Adaptive Immune Receptor Repertoire Data.

High-throughput sequencing (HTS) of immunoglobulin (B-cell receptor, antibody) and T-cell receptor repertoires has increased dramatically since the technique was introduced in 2009 (1-3). This experimental approach explores the maturation of the adaptive immune system and its response to antigens, pathogens, and disease conditions in exquisite detail. It holds significant promise for diagnostic and therapy-guiding applications.

Novel Approaches to Analyze Immunoglobulin Repertoires.

Analysis of immunoglobulin (Ig) repertoires aims to comprehend Ig diversity with the goal of predicting humoral immune responses in the context of infection, vaccination, autoimmunity, and malignancies. The first next-generation sequencing (NGS) analyses of bulk B cell populations dramatically advanced sampling depth over previous low-throughput single-cell-based protocols, albeit at the expense of accuracy and loss of chain-pairing information. In recent years the field has substantially differentiated, with bulk analyses becoming more accurate while single-cell approaches have gained in throughput.

sciReptor: analysis of single-cell level immunoglobulin repertoires.

Background: The sequencing of immunoglobulin (Ig) transcripts from single B cells yields essential information about Ig heavy:light chain pairing, which is lost in conventional bulk sequencing experiments. The previously limited throughput of single-cell approaches has recently been overcome by the introduction of multiple next-generation sequencing (NGS)-based platforms. Furthermore, single-cell techniques allow the assignment of additional data types (e.

The promise and challenge of high-throughput sequencing of the antibody repertoire.

Efforts to determine the antibody repertoire encoded by B cells in the blood or lymphoid organs using high-throughput DNA sequencing technologies have been advancing at an extremely rapid pace and are transforming our understanding of humoral immune responses. Information gained from high-throughput DNA sequencing of immunoglobulin genes (Ig-seq) 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. Rapid progress in the development of experimental protocols and informatics analysis tools is helping to reduce sequencing artifacts, to achieve more precise quantification of clonal diversity and to extract the most pertinent biological information.