Genomic Precision

 A SeraCare blog focused on precision medicine and advanced clinical diagnostics

This is the Number One Risk to Your Clinical Sequencing Assay

Critical missteps during the assay development phase can cause expensive delays and risk the quality of an assay. How can you be sure your bioinformatics pipeline is correctly calling variants?

Posted by Trevor Brown on Oct 17, 2017 11:00:00 AM

If you’re relying on remnant patient samples to tell you how well your lab's bioinformatics pipeline can call clinically important variants, you might be missing more than you realize.

In our experience, the bioinformatics pipeline can be the weakest link in assay development for many labs. Just because a variant is sequenced correctly doesn’t always mean that it will be called. And false-positives are just as bad.

  • Sometimes it’s an issue of allele frequency. For example, we’ve seen cases where labs could detect certain mutations at 10% allele frequency, but as soon as the frequency dropped to 7%, they stopped detecting it.
  • Other cases are caused by the complexity of the variant. For example, even at low allele frequencies, a lab may pick up relatively easy-to-detect single-nucleotide variants (SNVs) but can have problems with insertion/deletion (INDEL) calling errors.

In both examples, the mutations aren’t missed because of sequencing or library preparation problems. As we’ve witnessed time and time again, when labs optimize their bioinformatics pipelines, they start picking up the low-frequency and difficult-to-detect variants again.

The catch is, you first have to know you’re missing something. In assay development, what you don’t know can seriously weaken your test.

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Topics: allele frequencies, next-generation sequencing, cancer profiling, bioinformatics

Multiplexed Reference Materials as Controls for Cardiomyopathy Diagnostic Next-Generation Sequencing

A Journal for Molecular Diagnostics article describing ‘an attractive addition to the repertoire of materials for the development, validation, and quality monitoring of clinical NGS assays’

Posted by Dale Yuzuki on Dec 13, 2016 11:01:00 AM

Hypertrophic Cardiomyopathy (HCM) is a disease where the heart muscle is enlarged and a significant cause of sudden cardiac death, and is frequently asymptomatic.  HCM is commonly caused by a mutation in one of nine heart muscle genes that comprise the sarcomere, and occurs at a prevalence of about 1 in 500 in the general population. HCM is the leading cause of cardiac death in young athletes in the United States.

Clinical genetic testing for mutations in the HCM-related genes has been ongoing for over a decade;  the GeneTest.org database reveals 105 laboratories offering some version of genetic testing.  While knowledge of prevalent pathogenic variants are available, the majority of variants remain private (that is, unpublished and not widely available). The move to NGS-based gene panels for HCM testing has lead to new challenges for test development, validation and routine quality control due to the inherent scarcity of samples, the cost of including numerous single mutations from these individual samples, and the lack of these materials for laboratories without a long history of testing.

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Topics: NGS assays, next gen sequencing, allele frequencies, Controls, Cardiomyopathy, DNA, Clinical Genetic Testing, multiplexed referenence materials, multiplexing

Reference Materials for Your Unique Reproducibility Needs

The distinction between accuracy and precision.

Posted by Matt Ryder on Oct 12, 2016 11:06:00 AM

If you took a university introductory statistics course, you may have learned the distinction between accuracy and precision.  It may likely have been presented with an archery analogy, where ‘Accurate’ was represented by arrows loosely clustered around the target’s bull’s-eye, ‘Precise’ was shown as a tight grouping displaced from the center, and ‘Accurate and Precise’ was depicted as what every archer aims for, a tight grouping directly at the bull’s-eye.  Suddenly, words that are used interchangeably in everyday conversation took on dramatically different meanings.


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Topics: NGS, Precision Medicine, FDA, Reference Material, SeraSeq, NGS assays, Tumor Mutation Mix, Sequencing quality control, Good Manufacturing Practices, variants, allele frequencies

 
 

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