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Rapid and flexible analytical characterization of oligonucleotides
Therapeutic oligonucleotides (ASOs, siRNA and conjugates) present analytical challenges due to their structural diversity and chemical modifications. In early discovery, robust UV×MS purity assessment and impurity profiling are essential for both crude oligonucleotide libraries prior to early screening and purified batches prior to advanced assays.
Here, we present a streamlined LC–UV–MS platform comprising two methods paired with (semi-)automated data processing to increase analytical throughput and accelerate decision-making. First, a flexible characterization method provides chromatographic separation and UV×MS purity/impurity analysis for purified ASOs, siRNA and conjugates, with semi-automated processing and automated reporting enabled by the Byos Oligo App. Second, a high-throughput method delivers online desalting, rapid MS fingerprinting, purity analysis and impurity-class profiling for crude ASO and siRNA single‑strand libraries at scale (hundreds per day).
Together, these methods provide a fast, scalable and fit‑for‑purpose framework for oligonucleotide characterization in early drug discovery.
Key takeaways:
- Fit‑for‑purpose LC–UV–MS workflows We developed two analytical methods, one for detailed characterization of purified batches and one for high‑throughput screening of crude libraries, covering early discovery needs.
- (Semi-)automated data analysis We integrated data analysis tools, such as the Byos Oligo workflow, to enable semi‑automated UV×MS purity determination and impurity assignment with automated reporting, reducing manual effort and variability.
- Standardized reporting We implemented standardized outputs to deliver consistent purity/impurity metrics for oligonucleotides, improving comparability across libraries and individual samples.
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