Development of a Multi-Variant Frequency Ladder for NGS Assay Workflows - ASHG 2014

Increasing adoption of NGS has shed light on the need for more standardized controls to evaluate and optimize system performance. Samples containing mutations of interest are difficult to source and cell line pooling experiments to determine limit of detection require significant investments of time and money. To simultaneously evaluate variant calling performance in over 200 unique amplicons across 50 genes targeted by NGS tests, AcroMetrix® has developed a proprietary genomic/synthetic DNA material containing over 550 mutations as a mixture of SNV’s indels and MNP’s. The limit of detection was then determined for 400+ variants using multiple platforms. Tumor samples were diluted with matched normal samples to mimic a range of frequencies. Linearity between the material and diluted tumor tissue samples were compared. Overall, highly multiplex controls with tunable frequencies allow for much more extensive, yet streamlined, assay evaluation and facilitate implementation and impart confidence to NGS testing.