TCRs are cell specific and represent a sort of "molecular tag" of T cells and have been widely studied to monitor the dynamics of T cells in terms of clonality and diversity in several contexts including lymphoid malignancies, infectious diseases, autoimmune diseases, and tumor immunology. This ability is achieved during thymic development through a complex molecular mechanism based on somatic recombination that leads to the expression of a very heterogeneous population of surface antigen receptors, the T Cell Receptors (TCRs). Using an in silico meta-repertoire generated from 108 replicates, we found that one genomic DNA-based method and two non-unique molecular identifier (UMI) RNA-based methods were more sensitive than UMI methods in detecting rare clonotypes, despite the better clonotype quantification accuracy of the latter.The peculiarity of T cell is their ability to recognize an infinite range of self and foreign antigens. Results from the 5' RACE-PCR methods were consistent among themselves but differed from the RNA-based multiplex-PCR results. Low RNA input generated non-representative repertoires. Most methods showed a lower ability to capture TRA than TRB diversity. We found marked differences in accuracy and intra- and inter-method reproducibility for T cell receptor α (TRA) and T cell receptor β (TRB) TCR chains. In this study, we systematically compared the results of nine commercial and academic TCRseq methods, including six rapid amplification of complementary DNA ends (RACE)-polymerase chain reaction (PCR) and three multiplex-PCR approaches, when applied to the same T cell sample. the T cell receptor (TCR) repertoire in health and disease can provide key insights into adaptive immune responses, but the accuracy of current TCR sequencing (TCRseq) methods is unclear. 14 AP-HP, Hôpital Pitié-Salpêtrière, Biotherapy (CIC-BTi) and Inflammation-Immunopathology-Biotherapy Department (i2B), Paris, France. 13 Sorbonne Université, INSERM, Immunology-Immunopathology-Immunotherapy (i3), Paris, France.12 Vaccine Research Center, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Bethesda, MD, USA.10 Division of Cardiology, Medical University of Graz, Graz, Austria.9 Diagnostic and Research Institute of Pathology, Medical University of Graz, Graz, Austria.8 DFG-Centre for Regenerative Therapies Dresden, Faculty of Medicine Carl Gustav Carus, Technische Universität Dresden, Dresden, Germany.7 Division of Infection and Immunity, University College London, London, UK.6 Adaptive Immunity Group, Central European Institute of Technology, Brno, Czechia.5 Center for Precision Genome Editing and Genetic Technologies for Biomedicine, Pirogov Russian National Research Medical University, Moscow, Russia.4 Genomics of Adaptive Immunity Department, Shemyakin and Ovchinnikov Institute of Bioorganic Chemistry, Moscow, Russia.3 Center of Life Sciences, Skoltech, Moscow, Russia.2 AP-HP, Hôpital Pitié-Salpêtrière, Biotherapy (CIC-BTi) and Inflammation-Immunopathology-Biotherapy Department (i2B), Paris, France.1 Sorbonne Université, INSERM, Immunology-Immunopathology-Immunotherapy (i3), Paris, France.
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