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Abstract:
Recognition of foreign- and self-antigens by T cells and its receptors
(TCRs) is key to adaptive immunity. To achieve effective immunity,
millions of T cells must be able to match any given antigen through their
unique, heterodimeric TCRs. This makes dissecting entire TCR repertoires
challenging: conventional methods fail to pair both TCR chains, thus
providing incomplete clonotype information. Further, single-cell
transcriptome approaches are costly and only capture a small fraction (<1%)
of the effective repertoire. Here we present our new method CITR-Seq
(combinatorial indexing T cell receptor sequencing), which identifies TCR
α and β pairings in hundreds of thousands of T cells from one individual.
CITR-Seq combines two features: multiplexed TCR transcript amplification
and single-cell combinatorial indexing. We bypass the need of specialized
equipment, enabling processing of more cells and samples than before.
To test our approach we construct TCR repertoires of CD8+ T cells isolated
from mouse spleen. Across samples we can consistently recover > 400,000
T cells with up to 70% successful TCR α and β pairing. By contrast, using
the commercial 10X Genomic’s Chromium Next GEM platform we were
only able to confidently recover clonotypes of fewer than 10,000 cells per
sample, thus clearly showing the boost in throughput. Next we applied
CITR-Seq to mouse sister-species (Mus castaneus and Mus spretus). We
can consistently recover diverged and evolved T cell repertoires confirming
CITR-Seq’s ability to better capture TCR diversity.
The method presented here greatly expands the scope of TCR repertoire
studies, allowing for the generation of population-scale repertoire profiles.
In the future, we expect CITR-Seq to be used to better understand the
dynamics of TCRs, e.g. the role between public and private TCR molecules.
