A Good NGS Panel Starts with a Good Probe: Celemics’ Probe Design Principles
The performance of a targeted sequencing panel ultimately comes down to the design of each individual probe. No matter how refined a wet-lab protocol may be, if the probes fail to properly capture the target regions or produce uneven coverage, the resulting data becomes difficult to trust. To address this, Celemics applies a consistent set of principles across every stage of panel design — from the choice of probe material, to how performance is validated, to how the results are ultimately reported.
Why RNA Probes
Celemics uses RNA as the probe material for its hybrid capture panels. RNA-DNA hybrids have higher binding stability (Tm) than DNA-DNA hybrids, which allows for higher stringency during the post-capture wash steps. This means non-specifically bound, off-target sequences can be removed more effectively while the intended target sequences remain stably bound — ultimately improving both on-target rate and specificity.
2X Tiling: Covering the Entire Target Region Through Probe Complementarity
To ensure complete coverage of target regions without gaps, Celemics applies a 2X tiling strategy. At least two independent probes — each with a distinct sequence composition — are designed to overlap across a given target region. This way, if one probe shows relatively lower efficiency due to GC content or sequence-specific secondary structure, another probe covering the same region can compensate for it. This complementary probe structure prevents a localized drop in performance from one probe from turning into a gap in overall coverage. Building on this baseline principle, Celemics also applies design know-how accumulated through years of designing and testing a wide range of custom panels — fine-tuning probe spacing and overlap positions for specific regions where this experience has shown it improves results.
Introducing Modifications to Ensure Probes Work as Intended
Sequence information alone is not enough to ensure a probe functions exactly as designed. A range of modifications addressing probe stability, suppression of non-specific binding, and resistance to degradation must also be applied. Celemics incorporates these modifications from the design stage onward, so that probes bind to their targets as intended and remain stable throughout the actual experimental process.
Probe Rebalancing Based on Real Wet-Lab Data
A common pitfall in panel design is relying solely on in silico predictions to estimate probe efficiency. In practice, however, probe behavior in a wet-lab environment is influenced by many variables — sequence characteristics, secondary structure, interactions with neighboring sequences — so there is always some gap between predicted and actual performance. Celemics provides customers with real experimental results so they can see realistic, actual performance figures, and performs rebalancing based on this data when needed. Rather than relying on simulated predictions, probe concentrations for underperforming regions are adjusted based on measured coverage data, resulting in a panel grounded in real-world performance.
A Transparent Panel Design Policy: Not Masking Hard-to-Capture Regions
Regions that are inherently difficult to capture due to their sequence characteristics — hard-to-capture regions — can exist in any panel. Excluding such regions from the design stage can artificially inflate the overall performance metrics that get reported. Celemics avoids this approach. As a principle, target regions requested by customers are included in the panel to the greatest extent possible, even when they are difficult to capture, and the resulting real-world performance is reported as is. Rather than trimming target regions to make the numbers look better, Celemics believes that faithfully including the regions customers need and sharing performance transparently is what builds a more trustworthy panel.
Summary
The choice of RNA as the probe material, structural coverage secured through 2X tiling, modifications that enhance probe stability, and rebalancing grounded in real data — all of these efforts point toward a single goal: capturing the target regions customers request, completely and reliably, with data they can trust. Celemics is committed to reporting honest performance metrics, while continuing to design and improve panels in pursuit of the highest possible capture efficiency.
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