Next Generation Sequencing (NGS) Internal Controls for Adventitious Agent Testing to Ensure Sensitivity for All Targets in Every Sample

Our proposal will develop a test kit—AccuKit— that streamline adventitious agent screening while directly ensuring every target is measured with the required sensitivity.

Project status

100% Completed

Industry Need

Adventitious agent testing of Master and Working cell banks is one critical aspect of purity testing as it minimizes risk of upstream contamination.
Adventitious agent testing is moving away from the use of animal models to more in vitro tests such as cell culture and PCR-based methods.

Solution

AccuGenomics and NC State will develop a test kit (“AccuKit”) that will streamline adventitious agent screening while directly ensuring every target is measured with the required sensitivity.

AccuKits will incorporate PCR primers specific for 22 known adventitious viruses and bacteria for high throughput NGS screening assays.
The kit will contain a mixture of competitive templates for every adventitious agent spiked into every sample as internal sensitivity controls; the expected test result for contamination should be positive for every spike-in target and negative for the native targets.
The platform allows for additional primer sets to be added to the AccuKit to meet future adventitious agent detection needs. The team intends to advance its MRL-4 level technology to MRL-6.

Impacts

Developed a demonstration test kit—AccuKit—that uses Internal Standards (IS) to ensure every target measured by NGS had the required sensitivity

Created a sensitive and rapid NGS library preparation for at least 15 agents

Developed a manufacturing SOP to produce the resulting AA AccuKit.

Further leveraged approach to create a COVID-19 AccuKit for customers using SNAQ controls

AccuKit will help Improve the testing of biopharmaceutical products by streamlining and shortening adventitious agent screening in Master and Working cell banks, minimizing risk of upstream contamination; increase sample throughput and decrease costs for biopharmaceutical safety testing; and improve the efficiency of adventitious agent detection by providing the missing quality controls to ensure a sensitive biosafety testing package for any biopharmaceutical product

Publications

Cleveland, M. H., Anekella, B., Brewer, M., Chin, P., Couch, H., Delwart, E., Huggett, J., Jackson, S., Martin, J., Monpoeho, S., Morrison, T., Ng, S. H. S., Ussery, D., & Khan, A. S. (2020). Report of the 2019 NIST-FDA workshop on standards for next generation sequencing detection of viral adventitious agents in biologics and biomanufacturing. Biologicals : Journal of the International Association of Biological Standardization, 64, 76–82. https://doi.org/10.1016/j.biologicals.2020.02.003

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Project Lead

Accugenomics Inc

Participating Organizations

North Carolina State University