SwabExpress: An End-to-End Protocol for Extraction-Free COVID-19 Testing

Abstract

Background The urgent need for massively scaled clinical testing for SARS-CoV-2, along with global shortages of critical reagents and supplies, has necessitated development of streamlined laboratory testing protocols. Conventional nucleic acid testing for SARS-CoV-2 involves collection of a clinical specimen with a nasopharyngeal swab in transport medium, nucleic acid extraction, and quantitative reverse transcription PCR (RT-qPCR) (1). As testing has scaled across the world, the global supply chain has buckled, rendering testing reagents and materials scarce (2). To address shortages, we developed SwabExpress, an end-to-end protocol developed to employ mass produced anterior nares swabs and bypass the requirement for transport media and nucleic acid extraction. Methods We evaluated anterior nares swabs, transported dry and eluted in low-TE buffer as a direct-to-RT-qPCR alternative to extraction-dependent viral transport media. We validated our protocol of using heat treatment for viral inactivation and added a proteinase K digestion step to reduce amplification interference. We tested this protocol across archived and prospectively collected swab specimens to fine-tune test performance. Results After optimization, SwabExpress has a low limit of detection at 2-4 molecules/uL, 100% sensitivity, and 99.4% specificity when compared side-by-side with a traditional RT-qPCR protocol employing extraction. On real-world specimens, SwabExpress outperforms an automated extraction system while simultaneously reducing cost and hands-on time. Conclusion SwabExpress is a simplified workflow that facilitates scaled testing for COVID-19 without sacrificing test performance. It may serve as a template for the simplification of PCR-based clinical laboratory tests, particularly in times of critical shortages during pandemics.

SARS-CoV-2 Epidemiology on a Public University Campus in Washington State

“…Samples collected at kiosks were transported to the Northwest Genomics Center at the University of Washington and tested for SARS-CoV-2 using a quantitative reverse transcription polymerase chain reaction (RT-qPCR) laboratory-developed test (LDT). The RT-qPCR consists of assays for 2 SARS-CoV-2 targets in duplicate and the human marker RNase P across 4 multiplexed reactions ( Supplementary Data ) [ 16 ]. A sample was considered positive if 3 or 4 replicates for RNase P had a cycle threshold (Ct) value <36 and SARS-CoV-2 had a value <40 ( Supplementary Data ).…”

“…Samples were defined as failed and considered “never tested” if RNase P was undetected in 2 or more reactions, or if there was a laboratory or operator error. Midstudy (November 18, 2020), we implemented an extraction-free testing method that yielded similar results, but Ct values from the 2 methods were not directly comparable [ 16 ]. Results were provided to participants through a research report hosted on a secure online portal that was accessed using a unique barcode identifier and date of birth.…”

Highly Sensitive Immunoresistive Sensor for Point-Of-Care Screening for COVID-19

“…The clinical testing lab was approved for SARS-CoV-2 screening assay based on RT-qPCR [36]. The Northwest Genomics Center (NWGC) SwabDirect SARS-CoV-2 detection assay was validated for the workflow of COVID-19 screening.…”

The Seattle Flu Study: when regulations hinder pandemic surveillance

“…Laboratory Improvement Amendments; and laboratory flexibility in response to new and emerging pathogens and supply-chain disruptions that may emerge 4 .…”