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Author Spotlight: Advancements in Multiplex Detection of Respiratory Viruses
Published on: November 10, 2023
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Multiplexing primer/probe sets for detection of SARS-CoV-2 by qRT-PCR
Garrett A Perchetti1, Arun K Nalla1, Meei-Li Huang1
1Department of Laboratory Medicine, Virology Division, University of Washington, Seattle, WA, United States.
Summary
Multiplexing assays for SARS-CoV-2 detection significantly improved laboratory throughput. This method accurately identified the virus while conserving valuable resources during the COVID-19 pandemic.
Area of Science:
- Clinical diagnostics
- Molecular biology
- Virology
Background:
- The COVID-19 pandemic caused global strain on healthcare and clinical laboratories.
- Limited resources and the need for broad diagnostic testing presented challenges.
- Early SARS-CoV-2 tests required parallel assays, consuming valuable reagents.
Purpose of the Study:
- To develop and evaluate a multiplexed assay for SARS-CoV-2 detection.
- To improve diagnostic efficiency and resource utilization in clinical laboratories.
Main Methods:
- Nasopharyngeal swabs were tested using a laboratory-developed test (LDT) based on CDC protocols.
- A triplex quantitative reverse transcriptase PCR assay was developed, combining two virus targets and an extraction control.
- Assay specificity was evaluated using samples with other respiratory infections.
Main Results:
- The triplex assay correctly identified SARS-CoV-2 in 98.4% of positive/inconclusive samples.
- All negative samples tested by single-plex LDT were also negative by triplexing.
- No cross-reaction was observed with other common respiratory pathogens.
Conclusions:
- Multiplexing two gene targets and an extraction control is comparable to parallel assays.
- This approach significantly improves assay throughput and conserves resources.
