Performance Testing

The following sections describe performance testing methods.

Analytical Performance Testing

Illumina tests the analytical performance of variant calling using an approach that covers the entire workflow including library preparation, sequencing, and secondary analysis. This approach is used to test a diverse selection of variants. When the variant calling pipeline is expanded to call a new variant class, this approach is always used.

The software may include results for beta features, which have not been fully evaluated for performance. For more information, refer to the release notes for your software version.

In Silico Characterization Methods

Illumina uses in silico testing to the test the ability of the software to call an expanded scope of clinically relevant variants, including rare variants. While Illumina has analytically verified the performance of the software for calling complex variants in EGFR, the in silico testing approach characterizes the feasibility of the software to call complex variants in other genes.

In in silico testing , variants of interest are extracted from public databases such as Cosmic and ClinVar. Each variant is simulated at different VAF levels by spiking in mutant reads into a normal FFPE background or by increasing or decreasing the coverage of exons in the normal FFPE sample (eg, to simulate exon-level CNVs). The simulated reads match the expected quality of typical FFPE samples, such as fragment length, error rate, and family size. After the simulation, the software processes samples with spiked-in variants and determines the results. This approach does not include library prep and sequencing of tumor FFPE samples that include the rare variants of interest. The software reports these variants, but analytical verification is not performed.