key: cord-0749804-hyp2qrn7
authors: ISHII, Toshiaki; SASAKI, Masakazu; YAMADA, Kageto; KATO, Daiki; OSUKA, Hiroyoshi; AOKI, Kotaro; MORITA, Toshisuke; ISHII, Yoshikazu; TATEDA, Kazuhiro
title: Immunochromatography and chemiluminescent enzyme immunoassay for COVID-19 diagnosis
date: 2021-02-25
journal: J Infect Chemother
DOI: 10.1016/j.jiac.2021.02.025
sha: d5bd31ec8a02ce0fbd51abb42913174dc4fe5931
doc_id: 749804
cord_uid: hyp2qrn7

Introduction The rapid and accurate detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is required to prevent the spread of COVID-19. This study evaluated the utility of two SARS-CoV-2 antigen detection methods. Methods We evaluated two types of antigen detection methods using immunochromatography (Espline) and quantitative chemiluminescent enzyme immunoassay (Lumipulse). RT-PCR was performed as a standard procedure for COVID-19 diagnosis. Lumipulse and RT-PCR were performed for all 486 nasopharyngeal swabs and 136 saliva samples, and the Espline test was performed for 271 nasopharyngeal swabs and 93 saliva samples. Results The sensitivity and specificity of the Espline test were 10/11 and 260/260 (100%), respectively for the nasopharyngeal swabs and 3/9 and 84/84 (100%), respectively for the saliva samples. High sensitivities for both saliva (8/9) and nasopharyngeal swabs (22/24) were observed in the Lumipulse test. The specificities of the Lumipulse test for nasopharyngeal swabs and saliva samples were 460/462 (99.6%) and 123/127 (96.9%), respectively. Conclusion The Espline test is not effective for saliva samples but is useful for simple and rapid COVID-19 tests using nasopharyngeal swabs because it does not require special devices. The Lumipulse test is a powerful high-throughput tool for COVID-19 diagnosis because it has high detection performance for nasopharyngeal swabs and saliva samples.

Coronavirus disease 2019 (COVID-19), which is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), is one of the most severe health concerns facing the world today [1] . A COVID-19 diagnosis is determined principally by detecting the viral genome RNA using a quantitative reverse transcription polymerase chain reaction (RT-PCR) in the patients' nasopharynx and saliva [1] . Although RT-PCR has high sensitivity and specificity, it remains difficult to perform in all clinical laboratories because it requires special devices and reagents. While antibody detection is also a useful tool for investigating past SARS-CoV-2 infection, this method may not be sufficient for the diagnosis of COVID-19 [2] . Antigen detection methods based on immunochromatography and fluorescence immunochromatography have been developed for the diagnosis of COVID-19 [3] . However, the sensitivity of immunochromatography was low (ca. 30.2-75.5%) and a fluorescence analyzer was required for fluorescence immunochromatography [3] .

More recently, a rapid antigen detection kit, Espline ® SARS-CoV-2 (Espline; Fujirebio Inc., Tokyo, Japan), was developed for the detection of the viral nucleocapsid antigen based on immunochromatography [4] . Additionally, a quantitative antigen detection reagent with high sensitivity and specificity, J o u r n a l P r e -p r o o f Lumipulse ® SARS-CoV-2 (Lumipulse; Fujirebio Inc.) based on a chemiluminescent enzyme immunoassay of nasopharyngeal swabs and saliva samples, was developed [5] . However, the information regarding these antigen detection tests for SARS-CoV-2 is limited, especially information with respect to the use of saliva samples. Therefore, this study evaluated the utility of two test was used for quantitative antigen detection on the Lumipulse G1200 system (Fujirebio) for 35 min. The Lumipulse test measurement range was from 0.6 to 5000 pg/mL. A previous report indicated that the correlation between the virus load determined using RT-PCR and antigen amount determined using the Lumipulse test was maintained up to the antigen amount of 10,000 pg/mL [6] ; thus, the antigen amount >5,000 pg/mL was not diluted in this study. According to the manufacturer's recommendations, the criteria for the Lumipulse test were Real-Time PCR System (Thermo Fisher Scientific) [7] . The viral RNA was extracted from the Lumipulse test suspension using the QIAamp Viral RNA Mini Kit (Qiagen, Hilden, Germany). A threshold cycle (Ct) value less than 35 was positive, and a Ct value of 35 or more was defined as below the limit of detection (LOD) [7] . Table 1 shows the results of each detection method. The Espline test for nasopharyngeal swabs was highly sensitive (10/11 were positive) and had high specificity (260/260, 100%), in addition to its high concordance with the RT-PCR results. In one false-negative result, the Ct value of the RT-PCR was 33.7 because of a low virus load. Conversely, the Espline test for saliva samples showed a significantly low sensitivity (3/9 were positive), suggesting that the Espline test was not suitable for saliva samples. According to previous reports on immunochromatography using nasopharyngeal swabs, the sensitivities were 30.2%-57.6% for the Respi-strip (CORIS BioConcept, Gembloux, Belgium) and 73.3%-75.5% for the Panbio COVID-19 Rapid Test Device (Abbot, Chicago, J o u r n a l P r e -p r o o f USA) [3, [8] [9] [10] . These previous reports on the diagnosis of COVID-19 using immunochromatography showed lower sensitivities than of our study. The high sensitivity and specificity of the Espline test may be related to the use of centrifugation to process the sample. However, the main purpose of centrifugation is to remove impurities and increase specificity, which may not significantly affect sensitivity. In addition, previous reports have indicated that high sensitivities were obtained during the first week or within nine days of symptom onset and with high viral loads [4, 8] . Therefore, the sensitivity of This is the first report to evaluate the usefulness of the Lumipulse test in diagnosing COVID-19 using saliva. Previous studies of the Lumipulse test using nasopharyngeal swabs indicated that the sensitivities and specificities were 55.2%-91.7% and 97.3%-99.6%, respectively [5, 11] , and that these differences might be dependent on the timing of the sampling mentioned above. The detailed results of the COVID-19 positive patients are shown in Table 2 . Almost all the samples used in this study were obtained under 7 days after onset ( Table   2 ). No association was found between the presence or absence of underlying diseases and testing results. The distribution of the SARS-CoV-2 antigen value in COVID-19 patients were 4.5 to >5000, and the geometric mean was 808.9 pg/mL. If positive results with low antigen value or indeterminate results are observed, it may be necessary to perform an additional test with using RT-PCR and to assess the patients' symptoms. There have been reports of a case of confusion to a false-positive result from the Lumipulse test [11] .

This study had a limitation. A limited number of positive SARS-CoV-2 samples (both nasopharyngeal swabs and saliva samples) were available for this study. Therefore, further evaluations of the Espline and Lumipulse tests might be needed.

In conclusion, although the Espline test could not be used for saliva samples, it is useful for a simple point-of-care testing because it does not require a special device. In contrast, although the Lumipulse test requires a special device and reagents, it is useful in routine diagnosis because the Lumipulse test has a performance similar to the RT-PCR and has a simpler procedure and higher throughput than RT-PCR.

TI helped in writing the original draft; he was also involved in the methodology and formal analysis of this paper. MS, KY, HO, DK, KA, TM, YI, and KT contributed to reviewing and editing and were also involved in the formal analysis and writing of the original draft.

J o u r n a l P r e -p r o o f 

World Health Organization declares global emergency: A review of the 2019 novel coronavirus (COVID-19)

SARS-CoV-2-IgG response is different in COVID-19 outpatients and asymptomatic contact persons

Evaluation of the PanBio COVID-19 rapid antigen detection test device for the screening of patients with COVID-19

Evaluation of clinical utility of novel coronavirus antigen detection reagent. Espline® SARS-CoV-2

Comparison of automated SARS-CoV-2 antigen test for covid-19 infection with quantitative RT-PCR using 313 nasopharyngeal swabs, including from seven serially followed patients

Clinical validation of quantitative SARS-CoV-2 antigen assays to estimate SARS-CoV-2 viral loads in nasopharyngeal swabs

Development of genetic diagnostic methods for detection for novel coronavirus 2019(ncov-2019) in Japan

PanBio antigen rapid test is reliable to diagnose SARS-CoV-2 infection in the first 7 days after the onset of symptoms

Development and potential usefulness of the COVID-19 Ag Respi-strip diagnostic assay in a pandemic context

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

Another false-positive problem for a SARS-CoV-2 antigen test in Japan

We would like to thank Editage for the English language editing of our manuscript.

Funding: This research did not receive any specific grant from funding agencies in the public, commercial, or not-for-profit sectors.J o u r n a l P r e -p r o o f