[HTML][HTML] Immune repertoire sequencing using molecular identifiers enables accurate clonality discovery and clone size quantification

KY Ma, C He, BS Wendel, CM Williams, J Xiao… - Frontiers in …, 2018 - frontiersin.org
KY Ma, C He, BS Wendel, CM Williams, J Xiao, H Yang, N Jiang
Frontiers in immunology, 2018frontiersin.org
Unique molecular identifiers (MIDs) have been demonstrated to effectively improve immune
repertoire sequencing (IR-seq) accuracy, especially to identify somatic hypermutations in
antibody repertoire sequencing. However, evaluating the sensitivity to detect rare T cells and
the degree of clonal expansion in IR-seq has been difficult due to the lack of knowledge of T
cell receptor (TCR) RNA molecule copy number and a generalized approach to estimate T
cell clone size from TCR RNA molecule quantification. This limited the application of TCR …
Unique molecular identifiers (MIDs) have been demonstrated to effectively improve immune repertoire sequencing (IR-seq) accuracy, especially to identify somatic hypermutations in antibody repertoire sequencing. However, evaluating the sensitivity to detect rare T cells and the degree of clonal expansion in IR-seq has been difficult due to the lack of knowledge of T cell receptor (TCR) RNA molecule copy number and a generalized approach to estimate T cell clone size from TCR RNA molecule quantification. This limited the application of TCR repertoire sequencing (TCR-seq) in clinical settings, such as detecting minimal residual disease in lymphoid malignancies after treatment, evaluating effectiveness of vaccination and assessing degree of infection. Here, we describe using an MID Clustering-based IR-Seq (MIDCIRS) method to quantitatively study TCR RNA molecule copy number and clonality in T cells. First, we demonstrated the necessity of performing MID sub-clustering to eliminate erroneous sequences. Further, we showed that MIDCIRS enables a sensitive detection of a single cell in as many as one million naïve T cells and an accurate estimation of the degree of T cell clonal expression. The demonstrated accuracy, sensitivity, and wide dynamic range of MIDCIRS TCR-seq provide foundations for future applications in both basic research and clinical settings.
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