- Design
- Assay development and validation, discovery, validation and clinical cohorts
- Population
- 144 glioma samples (52 discovery, 39 validation, 53 clinical)
- Primary outcome
- Concordance of NGS-inferred copy number changes with FISH
- Effect
- 100% concordance with FISH in validation; 90% fewer FISH studies in routine use
A Canadian molecular laboratory wrote a custom analysis for an amplicon-based sequencing panel to infer the chromosome-level copy number changes used in integrated glioma diagnosis: 1p/19q codeletion, gain of chromosome 7 with loss of 10, EGFR amplification and CDKN2A/B homozygous deletion.
Thresholds were set in 52 retrospective cases, then tested against FISH in 39 prospective cases, with complete agreement. Strict thresholds were set for full specificity, with a grey zone sending samples to reflex FISH to protect sensitivity. Samples with under 30% tumour were excluded. In 53 routine cases, the approach cut FISH requests by 90%, shortened turnaround and lowered cost.
The validation is small and specific to one panel and pipeline, so other laboratories would need their own validation. But the principle, getting sequence variants and copy number from one test, is attractive for laboratories where FISH capacity is limited. It was published in July 2026.
- Consider whether your existing NGS panel can be validated to call glioma copy number changes.
- Keep reflex FISH for results in the grey zone between thresholds.
- Record tumour cellularity; samples under 30% were excluded here.
- Validate locally before replacing FISH; these thresholds are specific to one assay and pipeline.
Why it matters
One test instead of several shortens the wait for an integrated glioma diagnosis.
Don't overread it
Complete agreement in 39 validation cases is encouraging but too few to establish accuracy across all tumour types.
The statistics, in plain English
Complete concordance in 39 cases means no disagreements were seen, but with that sample a small error rate could still exist. The thresholds were chosen to be conservative, sending uncertain cases to FISH.
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