Determining the limit of detection for SARS-CoV-2 Omicron variant using rapid antigen tests

In a recent study posted to the medRxiv* pre-print server, researchers examined the analytical sensitivity of three widely used rapid antigen tests (RATs) used in the USA to detect the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Omicron variant, compared with the WA1 strain.

Currently, three antigen tests are widely used in the United States – the Abbott Binax Now, the AccessBio CareStart, and the LumiraDx – to determine the limit of detection (LoD) with gamma-irradiated or heat-inactivated preparations of the USA WA1/2020 (WA1) reference strains which does not have mutations in nucleocapsid protein (N).

The U.S. Food and Drug Administration (FDA) issued emergency use authorization (EUA) for RATs as useful in the diagnosis and surveillance of SARS-CoV-2. These tests target a genetically-conserved N protein to detect SARS-CoV-2 variants in clinical samples.

Amid the ongoing coronavirus disease (COVID-19) pandemic, the rise of Omicron, the B.1.1.529 variant, compelled the scientific community to speculate that this new SARS-CoV-2 variant might compromise the performance of all widely used antigen tests, as it harbors P13L, Δ31- 33, R203K, and G204R mutations in its N protein.

About the study

In the present study, researchers used cultured plaque-titered live Omicron and WA1 strain of SARS-CoV-2 to assess differences in the LoD with the Binax, CareStart, and LumiraDx tests. The study reported LoD values in plaque-forming units (PFU)/mL, converted from TCID50/mL to PFU/mL by multiplying the TCID50/mL by 0.7, a standard conversion based on the Poisson distribution.

Ten-fold serial dilutions in phosphate-buffered saline (PBS) ranging from 2.5×10⁴ to 2.5 PFU/mL were applied to swabs in 50µL volumes and tested in triplicate according to manufacturer instructions. While Binax and CareStart kits contained all the needed consumables, iClean foam swabs were used with the LumiraDx test.

The researchers identified the lowest 10-fold dilution with three replicate positive tests, then tested three-fold dilutions at this concentration until the lowest dilution (the LoD), in which at least 19/20 replicates (95%+) were positive, was identified.

Findings

The LoD values for Omicron and WA1 as measured by LumiraDx, Binax, and CareStart were 2.5×10²; 8.3×10¹, and 2.5×10² PFU/mL; and 2.8×10³ and 8.3×10³ PFU/mL, respectively.

For each tested variant, PFU and genome copies (gc/swab) had an almost similar relationship indicating that the Omicron mutations did not significantly alter the underlying diagnostic relationships and parameters of all the RATs.

There was a slight deviation from the manufacturers’ LoDs, most likely introduced in the results as the researchers used a live virus, analyte volume, and swab type.

Conclusions 

Previous studies did not meet the FDA’s EUA standards where 20 LoD replicates were crucial or included tests unavailable in the United States. The present study findings were consistent with previous investigations evaluating RATs sensitivities, but were more effective in demonstrating the sensitivity of Binax, CareStart, and LumiraDx tests than previous studies. 

Together, the findings demonstrated that the RATs detected Omicron effectively, alleviating concerns about the impact of the N mutations on the sensitivity of currently used RATs. Thus, RATs will continue to be a critical public health tool for keeping the SARS-CoV-2 Omicron variant transmission under control.

*Important notice

medRxiv publishes preliminary scientific reports that are not peer-reviewed and, therefore, should not be regarded as conclusive, guide clinical practice/health-related behavior, or treated as established information.

Written by

Neha Mathur

Neha Mathur has a Master’s degree in Biotechnology and extensive experience in digital marketing. She is passionate about reading and music. When she is not working, Neha likes to cook and travel.