A high-throughput nanoscale imaging platform for direct visualization and rapid measurement of telomeres at single-molecule resolution
Abstract
Telomere length is a biomarker of aging, cancer and telomeropathies, with some of the strongest health-related associations occurring at the extremes of the telomere-length spectrum. Critically short telomeres can drive chromosome instability, cellular stress and senescence, yet conventional assays largely provide population-average measurements that obscure these biologically important molecules, and the underlying heterogeneity of individual chromosome ends. In addition, amplification bias associated with highly repetitive telomeric sequences and inter-laboratory variability can limit measurement reproducibility. Although emerging technologies provide higher-resolution analyses, they often require high DNA input, specialized sample preparation, or complex instrumentation and bioinformatic tools, restricting broader adoption. Here, we present a high-throughput single-molecule platform that overcomes these barriers through an automated, user-friendly nanoscale imaging workflow compatible with standard genomic DNA preparations. The method combines a unique dual CRISPR-Cas9 telomere boundary labeling strategy with amplification-free magnetic-bead-based enrichment, preserving the native telomere-length distribution while minimizing representation bias. Using only 500 ng of input DNA and 2–4 h of hands-on processing, the assay accurately resolves intact telomeres spanning ~50 bp to >50 kb within the same sample, enabling simultaneous characterization of the full telomere-length spectrum. It also demonstrates high reproducibility across technical replicates and independent operators (ICC up to 0.90). Validation in telomere-length models, replicative aging systems, and clinical biospecimens, together with subtelomeric barcoding for chromosome-arm assignment, establishes a scalable platform for chromosome-resolved telomere profiling, biomarker development, and future clinical implementation in aging, cancer, and telomere biology.
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BibTeX
@misc{jacksoncook2026highthroughput,
title = {{A high-throughput nanoscale imaging platform for direct visualization and rapid measurement of telomeres at single-molecule resolution}},
author = {Collen Jackson-Cook and Lilia Gheghiani and Andrey Mikheykin and Dylan Van Laeken and Daniel Guest and Kellie Archer and Harold Riethman and Jason Reed},
howpublished = {Research Square},
year = {2026},
month = oct,
doi = {10.21203/rs.3.rs-11283393/v1},
url = {https://doi.org/10.21203/rs.3.rs-11283393/v1},
note = {CC BY 4.0},
}
RIS
TY - UNPB
TI - A high-throughput nanoscale imaging platform for direct visualization and rapid measurement of telomeres at single-molecule resolution
AU - Collen Jackson-Cook
AU - Lilia Gheghiani
AU - Andrey Mikheykin
AU - Dylan Van Laeken
AU - Daniel Guest
AU - Kellie Archer
AU - Harold Riethman
AU - Jason Reed
PY - 2026
DA - 2026/10/28
PB - Research Square
DO - 10.21203/rs.3.rs-11283393/v1
UR - https://doi.org/10.21203/rs.3.rs-11283393/v1
AB - Telomere length is a biomarker of aging, cancer and telomeropathies, with some of the strongest health-related associations occurring at the extremes of the telomere-length spectrum. Critically short telomeres can drive chromosome instability, cellular stress and senescence, yet conventional assays largely provide population-average measurements that obscure these biologically important molecules, and the underlying heterogeneity of individual chromosome ends. In addition, amplification bias associated with highly repetitive telomeric sequences and inter-laboratory variability can limit measurement reproducibility. Although emerging technologies provide higher-resolution analyses, they often require high DNA input, specialized sample preparation, or complex instrumentation and bioinformatic tools, restricting broader adoption. Here, we present a high-throughput single-molecule platform that overcomes these barriers through an automated, user-friendly nanoscale imaging workflow compatible with standard genomic DNA preparations. The method combines a unique dual CRISPR-Cas9 telomere boundary labeling strategy with amplification-free magnetic-bead-based enrichment, preserving the native telomere-length distribution while minimizing representation bias. Using only 500 ng of input DNA and 2–4 h of hands-on processing, the assay accurately resolves intact telomeres spanning ~50 bp to >50 kb within the same sample, enabling simultaneous characterization of the full telomere-length spectrum. It also demonstrates high reproducibility across technical replicates and independent operators (ICC up to 0.90). Validation in telomere-length models, replicative aging systems, and clinical biospecimens, together with subtelomeric barcoding for chromosome-arm assignment, establishes a scalable platform for chromosome-resolved telomere profiling, biomarker development, and future clinical implementation in aging, cancer, and telomere biology.
SN - 2693-5015
ER -