key: cord-0777435-5si9rdby
authors: Orsi, Andrea; Pennati, Beatrice Marina; Bruzzone, Bianca; Ricucci, Valentina; Ferone, Diego; Barbera, Paolo; Arboscello, Eleonora; Dentone, Chiara; Icardi, Giancarlo
title: On-field evaluation of a ultra-rapid fluorescence immunoassay as a frontline test for SARS-COV-2 diagnostic
date: 2021-05-28
journal: J Virol Methods
DOI: 10.1016/j.jviromet.2021.114201
sha: 8f78034cc567f38e7f3c4f92a13576c91eabc4ad
doc_id: 777435
cord_uid: 5si9rdby

BACKGROUND: Viral RNA amplification by real-time RT-PCR still represents the gold standard for the detection of SARS-CoV-2, but the development of rapid, reliable and easy-to-perform diagnostic methods is crucial for public health, because of the need of shortening the time of result-reporting with a cost-efficient approach. OBJECTIVES: The aim of our research was to assess the performance of FREND(TM) COVID-19 Ag assay (NanoEntek, South Korea) as a ultra-rapid frontline test for SARS-COV-2 identification, in comparison with RT-PCR and another COVID-19 antigen fluorescence immunoassay (FIA). STUDY DESIGN: The qualitative FIA FREND(TM) test, designed to detect within 3 minutes the Nucleocapsid protein of SARS-CoV-2, was evaluated using nasopharyngeal swabs in Universal Transport Medium (UTM™, Copan Diagnostics Inc, US) from suspected COVID-19 cases who accessed the Emergency Room of the Ospedale Policlinico San Martino, Genoa, Liguria, Northwest Italy. Diagnostic accuracy was determined in comparison with SARS-CoV-2 RT-PCR and STANDARD F(TM) COVID-19 Ag FIA test (SD BIOSENSOR Inc., Republic of Korea). RESULTS: In November 2020, 110 nasopharyngeal samples were collected consecutively; 60 resulted RT-PCR positive. With respect to RT-PCR results, sensitivity and specificity of FREND(TM) COVID-19 Ag test were 93.3% (95% CI: 83.8-98.2) and 100% (95% CI: 92.9-100), respectively. FREND(TM) and STANDARD F(TM) COVID-19 Ag FIA assays showed a concordance of 96.4% (Cohen’s k = 0.93, 95% CI: 0.86-0.99). CONCLUSIONS: FREND(TM) FIA test showed high sensitivity and specificity in nasopharyngeal swabs. The assay has the potential to become an important tool for an ultra-rapid identification of SARS-COV-2 infection, particularly in situations with limited access to molecular diagnostics.

Objectives. The aim of our research was to assess the performance of FREND TM COVID-19 Ag assay (NanoEntek, South Korea) as a ultra-rapid frontline test for SARS-COV-2 identification, in comparison with RT-PCR and another COVID-19 antigen fluorescence immunoassay (FIA).

Study design. The qualitative FIA FREND TM test, designed to detect within 3 minutes the Nucleocapsid protein of SARS-CoV-2, was evaluated using nasopharyngeal swabs in Universal Transport Medium (UTM™, Copan Diagnostics Inc, US) from suspected COVID-19 cases who accessed the Emergency Room of the Ospedale Policlinico San Martino, Genoa, Liguria, Northwest Italy. Diagnostic accuracy was determined in comparison with SARS-CoV-2 RT-PCR and STANDARD F TM COVID-19 Ag FIA test (SD BIOSENSOR Inc., Republic of Korea).

Results. In November 2020, 110 nasopharyngeal samples were collected consecutively; 60 resulted RT-PCR positive. With respect to RT-PCR results, sensitivity and specificity of FREND TM COVID-19 Ag test were 93.3% (95% CI: 83.8-98.2) and 100% (95% CI: 92.9-100), respectively. FREND TM and STANDARD F TM COVID-19 Ag FIA assays showed a concordance of 96.4% (Cohen's k = 0.93, 95% CI: 0.86-0.99).

Conclusions. FREND TM FIA test showed high sensitivity and specificity in nasopharyngeal swabs.

The assay has the potential to become an important tool for an ultra-rapid identification of SARS-COV-2 infection, particularly in situations with limited access to molecular diagnostics. In 2020, the increasing molecular analysis demand due to the COVID-19 pandemic raised several critical issues such as requirement of special equipment, laboratory reagents and skilled staff.

J o u r n a l P r e -p r o o f Therefore, looking for alternative diagnostic solutions to implement a molecular screening strategy extended to a large number of subjects and to counter the SARS-COV-2 spread has become a priority for the health care systems [3] .

In the last months, several easy to perform rapid antigen detection tests were developed and recommended in some countries as first line laboratory strategy for COVID-19 diagnostic [4, 5] .

The aim of the present research was to assess the performance of FREND TM COVID-19 Ag assay (NanoEntek, South Korea) as a frontline test for SARS-COV-2 identification in comparison to the available molecular techniques and to the STANDARD F COVID-19 Ag FIA test (SD BIOSENSOR Inc., Republic of Korea), a rapid fluorescent immunoassay test currently used in clinical practice [6] .

In November 2020, at the regional reference laboratory for COVID-19 diagnostic located into In 

In the ongoing COVID-19 pandemic context, diagnostic testing for SARS-COV-2 is crucial in order to limit the spread of the virus as well as appropriately manage infected patients. Different diagnostic test manufacturers have developed rapid tests based on SARS-COV-2 proteins detection in respiratory samples. However, the analytical performances of these rapid antigenic tests depend on different factors including the viral load, the quality of the specimen and how it is processed.

In this study, we determined the performance characteristics of the FREND TM COVID-19 Ag test for detecting SARS-COV-2 virus in respiratory samples and compared the results with RT-qPCR as the gold standard and another FIA antigen test. Our data showed that FREND TM COVID-19 Ag proved to be more sensitive and had several advantages such as the rapid answer in 3 minutes and the non-requirement of special equipment or personnel skills compared with molecular techniques.

The number of swabs collected and tested in this study could be a limit for our research; moreover, the Ct values of positive samples were never higher than 35. However, the study was performed in a "real world" clinical setting and within a few days, in order to evaluate the possible use of FREND TM COVID-19 Ag as a screening tool in a large reference hospital.

Although negative results cannot rule out SARS-COV-2 infection and made this test of little help when it is necessary to evaluate the progress of the disease and therefore, the state of recovery of the patient, our data suggest that FREND TM system could be a useful device particularly in situations with limited access to molecular diagnostics and the need of rapid results in order to appropriately manage people and resources, i.e. Emergency Rooms or outpatient facilities.

In conclusion, FREND TM COVID-19 Ag assay represents a valid frontline test for COVID-19 screening and it could ease the burden on the laboratories, reducing the time spent for diagnosis and the use of RT-qPCR.

We thank Arrow Diagnostics Srl for the donation of the FREND TM COVID-19 Ag kits to pursue the study. No other specific grant from public funding agencies was received. 

The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

Detection of 2019 novel coronavirus (2019-nCoV) by real-time RT-PCR

Laboratory Testing Strategy Recommendations for COVID-19

Evaluation of rapid antigen test for detection of SARS-CoV-2 virus

Low performance of rapid antigen detection test as frontline testing for COVID-19 diagnosis

Evaluation of a novel antigen-based rapid detection test for the diagnosis of SARS-CoV-2 in respiratory samples

Urgent need of rapid tests for SARS CoV-2 antigen detection: Evaluation of the SD-Biosensor antigen test for SARS-CoV-2

Republic of Korea), compared with RT-qPCR results. Swabs were divided into 3 groups based on RT-qPCR Ct values for N gene (<26, 26-30, 31-35) and into 2 groups according time from the onset of symptoms to sample obtention

Swabs FREND TM COVID-19 Ag STANDARD F COVID-19 Ag FIA Sensitivity (%, 95% C.I.)

We thank all staff of the Hygiene Unit, Regional Reference Laboratory for COVID-19 diagnostic, for the technical assistance offered for the study conduction. We special thank Simona Boccotti,