Tumour Purity and Sequencing: How Much Cancer Is in the Sample

Why Purity Sets a Detection Ceiling

If only 20 percent of the cells in a sample are cancer, a heterozygous mutation present in every cancer cell will appear in roughly 10 percent of sequencing reads, because half of the tumour DNA copies carry it.

If the assay's limit of detection is around 5 percent, that mutation is still detectable; if purity drops to 10 percent, the same mutation falls near or below the detection threshold and can be missed.

How Purity Is Estimated

Pathologists estimate purity visually on a stained slide, but this is approximate and observer-dependent. Bioinformatic methods infer purity from the sequencing data itself, using the allele fractions of many variants and copy-number patterns.

The two estimates do not always agree, and low-purity samples are harder to analyse reliably by either method.

Consequences for Different Findings

Low purity most affects the detection of subclonal mutations, low-level copy-number changes and, especially, homozygous deletions, which can be masked by the normal-cell signal.

Tumour mutational burden and microsatellite instability estimates are also less reliable at low purity because there is less tumour signal to measure.

Improving the Sample

Macrodissection or microdissection, where the pathologist marks and the laboratory scrapes a tumour-rich area of the slide, raises effective purity. Choosing a more cellular block, or requesting a fresh biopsy, are other options.

Laboratories usually set a minimum purity threshold below which they will not report, or will report with a caveat.

Interpretation Notes

A report should state the estimated tumour content. A negative result from a low-purity sample is weak evidence of true absence.

If a clinically expected mutation is not found and purity is low, repeat testing on a better sample is often warranted before concluding the tumour is negative.

Key Takeaways

• ·Tumour purity is the fraction of cancer cells in a specimen and caps detection sensitivity.
• ·Low purity can push real mutations below the assay's limit of detection.
• ·Homozygous deletions and subclonal changes are the most affected.
• ·Macrodissection and sample selection can raise effective purity.