Analytical and clinical evaluation of antigen-based rapid diagnostic tests against the most encountered SARS-CoV-2 variants

By Dr. Priyom Bose, Ph.D.Reviewed by Sophia CoveneyAug 2 2023

A recent study posted on the medRxiv* preprint server has assessed the sensitivity of antigen-based rapid diagnostic tests (Ag-RDTs). 

*Important notice: medRxiv publishes preliminary scientific reports that are not peer-reviewed and, therefore, should not be regarded as conclusive, guide clinical practice/health-related behavior, or treated as established information.

Background

The rapid spread of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) led to the coronavirus disease 2019 (COVID-19) pandemic.

In order to combat this pandemic, it is extremely important to prevent further transmission of infection. To achieve this, it is imperative to identify SARS-CoV-2-infected individuals and isolate them from healthy persons quickly.

Ag-RDTs can be performed at home by lay individuals to detect an infection. These diagnostic tests are inexpensive, do not require skilled personnel, and provide results rapidly.

In the context of the COVID-19 pandemic, Ag-RDTs have played a crucial role in controlling disease transmission. However, the efficiency of these tests has been challenged by the emergence of new SARS-CoV-2 variants.

The undetected SARS-CoV-2 variants of concern (VOC) have particularly threatened to trigger new waves of global infections.

At present, the SARS-CoV-2 Omicron (B.1.1.529) variant, which has been classified as a VOC, is dominantly circulating in most countries across the world. A total of five Omicron sub-lineages, namely, BA.1, BA.2, BA.3, BA.4, and BA.5, have been identified as of March 2023.

The Omicron variant contains the highest number of mutations throughout the genome that include spike (S), envelope (E), membrane (M), and nucleocapsid (N) protein regions. 

Most Ag-RDTs have been developed targeting the N protein of the wild-type (WT) SARS-CoV-2 (ancestral strain). Preliminary analytical and clinical assessments of a small number of brands have presented contradictory data about the efficacy of existing Ag-RDTs against the Omicron variant. 

About the study

In this study, a total of 34 commercially available COVID-19 Ag-RDTs were assessed for the detection of the Omicron VOC (BA.1 and BA.5). Furthermore, this finding was compared with the sensitivity against Alpha, Gamma, and Delta VOCs and WT. 

All Ag-RDTs evaluated in this study were based on lateral flow assays (LFA). Here, thirty-one of the Ag-RDTs used colorimetric gold nanoparticle detection, two used fluorescence, and one used microfluidic immunofluorescence technology.

All clinical samples were obtained from the ‘Facilitating Accelerated Clinical Evaluation of Novel Diagnostic Tests for COVID-19’ (FALCON) study. Nasopharyngeal (NP) swabs were collected from symptomatic participants who visited the COVID-19 test center in Liverpool John Lennon Airport, UK, between January 2021 and March 2022. 

Study findings

Out of the 34 tests that were analyzed, 31 Ag-RDTs exhibited sensitivity for Omicron sub lineage BA.5. This analysis was based on plaque forming units per milliliter (pfu/mL), fulfilling the criteria set by the British Department of Health and Social Care (DHSC).

As per the recommendations of the World Health Organization (WHO), the limit of detection (LOD) of Omicron VOC BA.5 was significantly lower than the WT and the Alpha VOC. However, the LOD value was similar to the Gamma, Delta, and Omicron BA.1 VOCs.

Only 23 Ag-RDTs exhibited sensitivity for Omicron BA.1. Taken together, these tests exhibited a significantly lower LOD with Omicron BA.5 compared to all other VOCs and WT. This high sensitivity against BA.5 could be attributed to the fact that BA.5 does not have any lineage-defining mutations in the N gene.

Clinical evaluations were performed on five out of thirty-four Ag-RDT brands. Compared to all tested Ag-RDTs brands, Onsite exhibited the best overall sensitivity, while Hotgen consistently performed poorly.

Strengths and limitations

The current study has several strengths, including the comprehensive evaluation of the analytical sensitivity of 34 commercially available Ag-RDT brands. Furthermore, all assessments were based on both viral isolates and clinical specimens.

One of the key limitations of this study is the use of retrospective frozen specimens instead of fresh swabs, as suggested by the majority of Ag-RDT manufacturers.

However, to rectify the possibility of potential RNA degradation after a freeze-thaw cycle, SARS-CoV-2 RNA was re-evaluated by RT-PCR at the time of conducting the Ag-RDT test.

Another limitation of this study is that the LOD experiments for the WT, Alpha, and Gamma viral isolates were not repeated. The current study used data from previously conducted experiments using a similar protocol, and these findings were compared with the new data for the Omicron and Delta variants.

Conclusions

The current study highlighted similar, if not better, LODs for the Omicron variant compared to other non-Omicron VOCs. Notably, both analytical and clinical assessments revealed a decreased sensitivity of the tested Ag-RDTs for Delta VOC.

In the future, commercially available Ag-RDTs must be continually assessed, especially those recommended for home testing, for their sensitivity for all SARS-CoV-2 VOCs, including the Delta variant. 

*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.