Diagnostic Precision

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How am I going to test my assay? Should I use patient samples or biosynthetic materials?

Posted by Dan Brudzewsky on Jan 4, 2018

Assay development and optimization for clinical genetics is increasingly challenging. In an era of clinical genomics, new technologies and clinical utilities constantly call for newer and better performing assays. Having access to an abundant supply of relevant and reliable test material is critical for quick assay development and well-documented assay performance.


What is relevant and reliable test material?

Traditionally, genetic assay development has relied on access to patient samples. However, clinical utility has advanced towards multigene disorders and tumor-derived DNA,  making adequate patient sample sourcing cumbersome, if at all possible. Therefore, agile and timely assay development also relies on biosynthetic materials to complement patient samples and ensure robust assay performance, as seen with Roche’s FDA-approved EGFR ctDNA assay, where lack of sufficient clinical specimens necessitated use of contrived samples for reproducibility, assay stability, and analytical sensitivity assessment.

Absence of evidence is not evidence of absence

Trends for clinical genetic tests are toward large gene panels and highly sensitive NGS or digital PCR assays. Each trend represents its own challenges, but the root question is similar: How does one demonstrate assay performance across a broad range of variant types?

SeraCare’s highly multiplexed biosynthetic reference material isn’t only a faster and more cost-effective route to NGS-based assay development, it will also provide assurance of a low false-negative rate for each successfully detected variant. Uniquely, its isogenic background also enables estimation of the equally important false-positive rate. Access to well-defined and robust test material is particularly critical for reliably documenting limit of detection for assays intended for testing tumor-derived samples (e.g. DNA variant allele frequency as low as 0.125% or presence or absence of fusion RNA transcripts). For example, tumors harboring the MET exon 14 deletion have increased sensitivity to MET inhibitors, such as crizotinib. However, more than 100 somatic variants, including intronic mutations, result in alternative splicing of the MET, but the frequency of MET exon 14 skipping in lung cancer is about 4% and even rarer in brain cancer. Therefore, having access to rare, but actionable variants and fusion RNA transcripts such as MET exon 14 skipping is important for building and implementing better clinical genomics assays.

To learn more on how to develop a clinical NGS-based assay, without losing your mind or shirt, click here.

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Topics: NGS, Assay Development