Accurately characterizing and quantifying the antibody repertoire are vital to discovering antibodies that recognize specific antigens of interest, including virus-neutralizing antibodies ( 5– 7) and therapeutic antibodies ( 8, 9), guiding the development of vaccines ( 10), detecting B-cell malignancies with high sensitivity ( 11), and monitoring immune status ( 9). This diversity enables B cells to recognize and neutralize a wide range of antigens, particularly invading pathogens and autoantigens accumulated in the body ( 3, 4). During B cell development, somatic recombination of variable (V), diversity (D, for heavy chain only) and joining (J) gene segments, non-templated (N) or palindromic (P) addition or subtraction of nucleotides at the junctions, and class switch recombination (CSR) and somatic hypermutation (SHM) upon activation all contribute to the diversity of the antibody repertoire ( 1, 2). The entire set of antibodies within an individual or tissue constitutes a tremendously diverse antibody repertoire. An antibody can neutralize a pathogen by recognizing a unique component (antigen) of the pathogen via its fragment antigen-binding (Fab) variable region. The removal of these artifacts will provide an accurate assessment of antibody repertoires and benefit related studies, especially mAb discovery and antibody-guided vaccine design.Īntibodies (Abs), also known as immunoglobulins (Igs), are the most important component of humoral immunity. ![]() Tested by ultra-deep Rep-seq data, DUMPArts removed inter-sample chimeras, which cause artifactual shared clones and constitute approximately 15% of reads in the library, as well as intra-sample chimeras with erroneous SHMs and constituting approximately 20% of the reads, and corrected base errors and amplification biases by consensus building. Here, a novel approach named DUMPArts, which improves the accuracy of antibody repertoires by labeling each sample with dual barcodes and each molecule with dual unique molecular identifiers (UMIs) via minimal PCR amplification to remove artifacts, is developed. However, polymerase chain reaction (PCR) amplification introduces extensive artifacts including chimeras and nucleotide errors, leading to false discovery of antibodies and incorrect assessment of somatic hypermutations (SHMs) which subsequently mislead downstream investigations. Antibody repertoire sequencing (Rep-seq) has been widely used to reveal repertoire dynamics and to interrogate antibodies of interest at single nucleotide-level resolution.
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