Blood Processing Delays Expose a Stable CORE of the T-Cell Receptor Repertoire

Delays between blood collection and sample processing are usually treated as a nuisance in T-cell receptor sequencing studies. A new analysis of peripheral blood samples from 211 patients with newly diagnosed colorectal cancer turns that nuisance into an instrument. By comparing samples processed shortly after collection with samples subjected to extended processing delays, researchers used the variability in handling time as a structured technical perturbation, and in doing so defined what they call CORE clonotypes — COnsistent Repertoire Elements — amino-acid-identical CDR3β sequences detected in both processing groups.

The starting observation was itself consequential for anyone sequencing immune repertoires. Delayed processing was associated with reduced clonotype richness, a depletion of singleton clonotypes, and altered diversity distributions. In other words, prolonged handling preferentially removes the low-frequency end of the repertoire, the rare clonotypes that are often the most biologically interesting and the most vulnerable to technical loss. This indicates that repertoire measurements remain sensitive to pre-analytical variables, and that samples processed on different timelines may not yield the same detectable repertoire composition.

Within that framework, the team defined CORE clonotypes operationally: a clonotype qualified as a CORE element if its amino-acid CDR3β sequence appeared in both the promptly processed and the delayed groups. The definition is deliberately simple, but the resulting set proved to be far from a random residue. The researchers characterized CORE clonotypes across multiple analytical levels, including abundance distributions, V and J gene segment usage, amino-acid composition, nucleotide redundancy, codon usage, biochemical properties, and correlation-network structure.

One of the most notable results concerns abundance. CORE clonotypes were largely distinct from abundance-matched clonotype sets, indicating that reproducible detection across perturbation conditions cannot be explained simply by the fact that frequent clonotypes survive and rare ones do not. Something beyond raw frequency makes these sequences reliably recoverable, suggesting that the CORE compartment reflects a non-random organizational feature of the peripheral repertoire rather than a sampling artifact.

The nucleotide level told an equally intriguing story. While the amino-acid composition of CORE clonotypes stayed conserved across processing conditions, these clonotypes exhibited elevated synonymous sequence diversity, meaning that multiple distinct nucleotide sequences encode the same CDR3β amino-acid string. Following sequencing-depth normalization, the CORE set also showed reduced codon-usage divergence and narrower distributions across multiple biochemical properties relative to clonotypes found in only one processing group. The convergent picture is one of constraint: the reproducible portion of the repertoire occupies a tighter region of sequence space than the portion that appears and disappears with handling time.

To probe whether these elements carry antigenic meaning, the researchers performed correlation-network analysis and identified a small CORE-associated subnetwork that was enriched for previously annotated antigen associations compared with randomized controls. The authors present this as an enrichment finding within their own dataset rather than proof of any specific immune response, but it suggests that the reproducible core is not molecularly anonymous: it contains clonotypes whose sequences have been linked to known antigens in earlier annotations.

The broader lesson of the study is methodological as much as biological. Pre-analytical variability, usually feared as a confounder in high-throughput immune profiling, can serve as a deliberate probe of repertoire structure. By asking which clonotypes withstand a reproducible technical stress, the researchers isolated elements that are non-randomly organized across abundance, gene usage, codon and protein-property levels. The work characterizes the peripheral TCRβ repertoire of a defined colorectal cancer cohort under controlled sequencing conditions, and the authors frame the CORE concept strictly as an operational definition grounded in processing-delay comparisons. For immune repertoire sequencing more generally, the message is that the reproducible skeleton of the repertoire can be exposed by subjecting it to perturbation — and that part of what looks like sequencing noise may, on closer inspection, outline repertoire structure itself.

Subject of Research: Reproducible structural components of the peripheral TCR repertoire revealed by blood processing delays

Article Title: CORE clonotypes: a reproducible structural component of the peripheral TCR repertoire revealed through variation in blood processing times

Article References: Bavly, O., Zilberberg, A., & Efroni, S. (2026). CORE clonotypes: a reproducible structural component of the peripheral TCR repertoire revealed through variation in blood processing times. BMC Bioinformatics. https://doi.org/10.1186/s12859-026-06715-6

Image Credits: AI Generated

DOI: 10.1186/s12859-026-06715-6

Keywords: TCR repertoire sequencing, CORE clonotypes, colorectal cancer, pre-analytical variability, CDR3β, codon usage, clonotype richness, immune profiling, bioinformatics, peripheral blood, reproducibility, correlation networks