期刊
NATURE BIOTECHNOLOGY
卷 39, 期 2, 页码 -出版社
NATURE PORTFOLIO
DOI: 10.1038/s41587-020-0656-3
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资金
- ERC-Advanced TRiPoD grant [322856]
- LabEx Transimmunom grant [ANR-11-IDEX-0004-02]
- RHU iMAP grant [ANR-16-RHUS-0001]
- European Research Area Network-Cardiovascular Diseases grant [ANR-18-ECVD-0001]
- iReceptorPlus (H2020 Research and Innovation Programme) grant [825821]
- Ministry of Science and Higher Education of the Russian Federation [075-15-2019-1789]
- National Institute of Allergy and Infectious Diseases
- National Institute for Health Research UCL Hospitals Biomedical Research
- DFG CRTD [FZ 111]
- Ministry of Education, Youth and Sports of the Czech Republic under project CEITEC 2020 [LQ1601]
- European Research Area Network-Cardiovascular Diseases (JCT2018) program
- European Research Area Network-Cardiovascular Diseases (AIR-MI Consortium) program
- European Research Area Network-Cardiovascular Diseases (JCT2018) grant
- H2020 Societal Challenges Programme [825821] Funding Source: H2020 Societal Challenges Programme
- Agence Nationale de la Recherche (ANR) [ANR-18-ECVD-0001] Funding Source: Agence Nationale de la Recherche (ANR)
- European Research Council (ERC) [322856] Funding Source: European Research Council (ERC)
This study systematically compared the results of nine different TCR sequencing methods on the same T cell sample, revealing significant differences in accuracy and reproducibility, with most methods showing a lower ability to capture TRA diversity and low RNA input affecting representativeness. Despite better clonotype quantification accuracy, some methods were found to be more sensitive in detecting rare clonotypes.
Monitoring 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. 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. We found marked differences in accuracy and intra- and inter-method reproducibility for T cell receptor alpha (TRA) and T cell receptor beta (TRB) TCR chains. Most methods showed a lower ability to capture TRA than TRB diversity. Low RNA input generated non-representative repertoires. Results from the 5 ' RACE-PCR methods were consistent among themselves but differed from the RNA-based multiplex-PCR results. 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. A comparison of T cell receptor repertoire profiling methods shows substantial differences in their outputs.
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