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American Journal of Infectious Diseases
Research Notes
Evaluation of A Rapid IgM-IgG Combined Antibody Test for
SARS-CoV-2 Infection: Single Italian Center Study
1Katia Margiotti, 2Marina Cupellaro, 2Sabrina Emili, 1Alvaro Mesoraca and 1,2,3Claudio Giorlandino
1Human Genetics Lab, Altamedica Main Centre, Viale Liegi 45, 00198 Rome, Italy
2Department of Biochemistry, Altamedica Main Centre, Viale Liegi 45, 00198 Rome, Italy
3Department of Prenatal Diagnosis, Altamedica, Fetal-Maternal Medical Centre, Viale Liegi 45, 00198 Rome, Italy
Article history
Received: 01-04-2020
Revised: 22-05-2020
Accepted: 20-06-2020
Corresponding Author:
Katia Margiotti
Human Genetics Lab,
Altamedica Main Centre,
Viale Liegi 45,
00198 Rome, Italy
Tel: +39 06 8505805
Fax: +39 068505815
Email: katia.margiotti@artemisia.it
Abstract: The need for timely establishment of diagnostic assays of Severe
Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) is demanded in
laboratories worldwide. We evaluated the performance of a flow
immunoassay which can detect IgM an IgG antibodies simultaneously
against SARS-CoV-2 virus in human blood within 15 min. Among the 132
positive novel Coronavirus Disease 19 (COVID-19) cases, 126 tests were
consistent with previous quantitative Real Time PCR (qRT-PCR) assays.
Among the 62 negative cases, 60 were consistent with qRT-PCR assays
except for 2 cases. In this study, 2019-nCOV/COVID-19 IgG/IgM Rapid
Test Device showed 95.5% sensitivity and 96.8% specificity. In conclusion,
Rapid IgM-IgG Combined Antibody Test showed high detection consistency
among all analysed samples. Suggesting that could be used for the rapid
screening of SARS-CoV-2 carriers, either symptomatic or asymptomatic.
Keywords: Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2),
Rapid IgM-IgG Combined Antibody Test, Novel Coronavirus Disease 19
(COVID-19)
Introduction
On December 31th, 2019 China reported first cases
of atypical pneumonia in Wuhan, the capital of Hubei
province. The causative virus was found to be a
betacoronavirus, closely related to the Severe Acute
Respiratory Syndrome Coronavirus (SARS-CoV-1) from
2003 and similar to Sarbecoviruses isolated from bats
(Wu et al., 2020; Zhou et al., 2020). It was therefore
termed SARS-CoV-2 and the disease it causes was
named Corona Virus Disease 2019 (COVID-19)
(CSGICTV, 2020). Several quantitative Real-Time RT-
PCR (qRT-PCT) protocols for detection of SARS-CoV-2
RNA have been developed and approved from Centers
for Disease Control and Prevention Nucleic acid in US
and are now widely employed to diagnose COVID-19
disease (Chu et al., 2020; Corman et al., 2020).
However, qRT-PCR take at least several hours to
complete and require certified laboratories, expensive
equipment and trained technicians to operate. Moreover,
these methods are dependent on the time‐window of
viral replication and they can potentially cause low
predictive rate results, thereby limiting the usefulness of
RT‐PCR in the field. Therefore, there is an urgent need
for additional tests, rapid and simple to use, to quickly
identify infected patients of SARS-CoV-2 virus,
especially by detecting IgM antibodies which are
observed about 12 day after infection, to prevent virus
transmission of infected patients (Infantino et al., 2020;
Okba et al., 2020; Zhao et al., 2020). However, it is
important to underline that the detection of
SARS‐CoV‐2 viral nucleic acid by RT‐PCR test is still
the current standard diagnostic method for COVID‐19.
At this time a great number of different rapid assays
have been proposed, but lack of analytical performance
and clinical validation are a major problem in terms of
reliability despite the easy access on the market of these
type of test (Rashid et al., 2020; Cassaniti et al., 2020).
The major type are serological assays based on colloidal
gold-labeled Immunochromatography (ICT) methods that
offer combination IgM and IgG detection (Rashid et al.,
2020). All these kits are based and use capture reaction
to detect SARS-CoV-2 IgM/IgG. For combination IgM
and IgG kit, the cassette has two detection bands (M and
G) and a quality control band (C). The M band is coated
with a monoclonal anti-human IgM antibody for
detecting SARS-CoV-2 antibody; the G line is fixed with
a reagent for detecting SARS-CoV-2 antibody; C line is
Katia Margiotti et al. / American Journal of Infectious Diseases 2020, 16 (2): 85.88
DOI: 10.3844/ajidsp.2020.85.88
86
fixed with a quality control antibody. All kits offer a
one-step method with results obtained within 15 min.
Samples that can be used are whole blood, serum or
plasma samples (Li et al., 2020). Recently, has been
reported that the detection accuracy of lateral flow
immunoassay anti-SARS-CoV-2 IgM and IgG antibodies
resulted in a sensitivity of 88.7% and a specificity of
90.6% (Li et al., 2020). The aim of this study was to
assess the diagnostic performance of a newly developed
lateral flow immunoassay anti-SARS-CoV-2 IgM and IgG
antibodies test, developed by using a combination of anti-
IgM-IgG Coronavirus 19 antibodies (2019-
nCOV/COVID-19 IgG/IgM Rapid Test Device,
Hangzhou Realy Tech Co., Ltd). Serological studies in
Italy and around the world appear to be still under
evaluation and reporting available laboratory data is
crucial in order to understand the utility of rapid antibody
detection during the course of SARS-CoV-2 infection.
Materials and Methods
Sample Collection
These samples were collected from various public
healthcare center and COVID-19 accredited healthcare
facilities in Italy, with oral consent from all participants
and approved by the local Ethics Committee of
Artemisia SPA. The 2019-nCOV/COVID-19 IgG/IgM
Rapid Test Device was conducted at Altamedica Medical
Centre (Rome, Italy) by clinical staffs who followed test
procedure described in the product inserts (2019-
nCOV/COVID-19 IgG/IgM Rapid Test Device,
Hangzhou Realy Tech Co., Ltd.)
Sample Testing
The IgM antibody and IgG antibody against SARS-
CoV-2 in blood samples were tested using 2019-
nCOV/COVID-19 IgG/IgM Rapid Test Device
(Hangzhou Realy Tech Co., Ltd) according to the
manufacturer’s instructions. These reagents are supplied
by Hangzhou Realy Tech Co., Ltd and resulted CE
marked and regularly registered with the Ministry of
Italian Health as an IVD Medical Device at N. 1923329.
Briefly, the pouched device was opened immediately
before use. Refrigerated blood samples used for the test,
are warmed to room temperature. During testing, 20 uL
whole blood sample are pipetted into the sample port
followed by adding 2 drops (about 20 uL) of dilution
buffer to drive capillary action along the strip. The entire
test took about 15 min to finish.
Data Analysis
The rapid SARS-CoV-2 IgG-IgM combined antibody
test kits were tested on blood samples coming several
hospitals and Italian COVID-19 accredited laboratories
in different provinces, with a total of 132 clinical
positive and 62 clinical negative patient blood samples.
The test performance was calculated with the Vassarstats
online calculator (http://www.vassarstats.net).
Results and Discussion
One hundred and thirty-two patients with qRT-PCR
positive SARS-CoV-2 and sixty-two qRT-PCR negative
SARS-CoV-2 infection were included in the study. No
clinical data were available along with the laboratory
results at that moment. The available characteristics of
the sample testing are reported in Table 1. All sample
were tested for viral antibody with a new 2019-
nCOV/COVID-19 IgG/IgM Rapid Test Device
(Hangzhou Realy Tech Co., Ltd). The sensitivity and
specificity of the rapid test newly developed were
verified in a total of 194 cases: 132 (positive) clinically
confirmed (by qRT-PCR test) SARS-CoV-2-infected
patients and 62 SARS-CoV-2 qRT-PCR negative cases.
In our study of the 132 SARS-CoV-2-infected patients,
126 resulted positive to the antibodies rapid test,
generating a sensitivity of 95.5% (CI95% 89.9-98.1), of
the 62 SARS-CoV-2 negative cases 2 tested positive,
generating a specificity of 95.8% (CI95% 87.8-99.4)
(Table 2). Moreover, the positive predictive value (PPV)
of antibodies test was 98.44% (126/128) and the
Negative Predictive Value (NPV) of antibodies test was
90.1% (60/66). It was also founded that 61.9% (78 out
of 126) of positive patients had both IgM and IgG
antibodies, while 7.9 % (10/126) where IgG positive
and 30% (38/126) where IgM positive. A singular IgM
response is an indication of a recent infection, while a
singular IgG response meaning that the infection was
encountered more than 2 months before the serological
test (Matricardi et al., 2020) (Table 2). Thus, the
antibody testing might play pivotal roles in the
following settings: (1) for suspected paucisymptomatic
patients, positive result of antibody increases the
confidence to make a COVID-19 diagnosis; (2) for
healthy subject, in this case of antibody positive result
the RNA should be tested more frequently and the close
contacts observed as well. Obviously, this test cannot
confirm virus presence, only provide evidence of recent
infection, but it provides an important immunological
evidence for physicians to make the exact diagnosis
along with other tests and to start treatment of patients.
Moreover, rapid laboratory diagnosis is essential for
commencement of infection control measures. Rapid
specific antigen tests have also been widely used in the
diagnosis of two other coronavirus infection disease,
Severe Acute Respiratory Syndrome (SARS) and
Middle East Respiratory Syndrome (MERS) (Lau et al.,
2004; Chen et al., 2016). Testing of specific antibodies
of SARS-CoV-2 in patient blood is a good choice for
rapid, simple, highly sensitive diagnosis of COVID-19.
Katia Margiotti et al. / American Journal of Infectious Diseases 2020, 16 (2): 85.88
DOI: 10.3844/ajidsp.2020.85.88
87
Table 1: Patient characteristics of the COVID-19 and control groups
Characteristics COVID-19 group
Age, years 35.5 (20.5-72.4)
Male sex 56 (42%)
Female sex 76 (58%)
Characteristics Control group
Age, years 32.1 (23.5-67.3)
Male sex 23 (37%)
Female sex 39 (63%)
Table 2: Detection sensitivity and specificity of 2019-nCOV/COVID-19 IgG/IgM rapid test device
qRT-PCR positive qRT-PCR negative
Sample Analysed 132 62
IgG&IgM Positive 78 0
IgG Positive 10 0
IgM Positive 38 2
Sensitivity 95.5% (CI95% 89.9-98.1)
Specificity 96.8% (CI95% 87.8-99.4)
There are some limitation to consider, like possible
cross-reactivity with other coronaviruses and flu viruses.
In fact, SARSCoV-2 belongs to betacoronavirus, in the
same family as SARS-CoV and MERS-CoV. Thus, there
is possibility of cross-reactivity with other coronaviruses
occurring and other bat-related SARS coronaviruses
remains to be clearly determined (Xiao et al., 2020).
Nevertheless, combination of nucleic acid qRT-PCR and
the IgM-IgG antibody test can provide more accurate
SARS-CoV-2 infection diagnosis. As today, only the
Cellex qSARS-CoV-2 IgG/IgM Rapid Test has been
validated by the US Food and Drug Administration (FDA)
in the Emergency Use Authorisation (EUA) category. The
objective of this study was to evaluate the overall clinical
performance and diagnostic value of a rapid serological
testing the 2019-nCOV/COVID-19 IgG/IgM Rapid Test
Device in detecting SARS-CoV-2-infected patients. Our
study represents a private clinical experience of the IgM-
IgG antibody test in an Italian laboratory for SARS-CoV-
2 antibodies detections. In our hands, the performance of
the 2019-nCOV/COVID-19 IgG/IgM Rapid Test Device
was comparable to that of recently published clinical
study, showing that viral serological testing is an effective
means of SARS-CoV-2 infection (Li et al., 2020).
Conclusion
A rapid 2019-nCOV/COVID-19 IgG/IgM Rapid Test
Device using lateral flow immune assay techniques was
evaluated. It takes less than 15 min to generate results
and determine whether there is recent SARS-CoV-2
infection. It is easy to use and no additional equipment is
required. Results from this study demonstrated that this
test is highly sensitive and specific. In conclusion, this
rapid test has great potential benefit for the fast screening
of SARS-CoV-2 infections and it has already generated
enormous interest in the medical community.
Author’s Contributions
Katia Margiotti: Provided insights on data analysis
and result and drafting the manuscript.
Marina Cupellaro and Sabrina Emili: Did the
laboratory analyses.
Alvaro Mesoraca and Claudio Giorlandino:
Conceived the study revised the manuscript.
All authors have approved the final article.
Ethics
The study was approved by the local Ethics
Committee of Artemisia SpA..
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