key: cord-254821-px4fe7mn
authors: Infantino, Maria; Grossi, Valentina; Lari, Barbara; Bambi, Riccardo; Perri, Alessandro; Manneschi, Matteo; Terenzi, Giovanni; Liotti, Irene; Ciotta, Giovanni; Taddei, Cristina; Benucci, Maurizio; Casprini, Patrizia; Veneziani, Francesca; Fabbri, Sergio; Pompetti, Adolfo; Manfredi, Mariangela
title: Diagnostic accuracy of an automated chemiluminescent immunoassay for anti‐SARS‐CoV‐2 IgM and IgG antibodies: an Italian experience
date: 2020-05-10
journal: J Med Virol
DOI: 10.1002/jmv.25932
sha: 
doc_id: 254821
cord_uid: px4fe7mn

A pandemic of coronavirus disease 2019 (COVID‐19) caused by severe acute respiratory syndrome coronavirus 2 (SARS‐CoV‐2) has been spreading throughout the world. Though molecular diagnostic tests are the gold standard for COVID‐19, serological testing is emerging as a potential surveillance tool, in addition to its complementary role in COVID‐19 diagnostics. Indubitably quantitative serological testing provides greater advantages than qualitative tests but today there is still little known about serological diagnostics and what the most appropriate role quantitative tests might play. Sixty‐one COVID‐19 patients and 64 patients from a control group were tested by iFlash1800 CLIA analyzer for anti‐SARS CoV‐2 antibodies IgM and IgG. All COVID‐19 patients were hospitalized in San Giovanni di Dio Hospital (Florence, Italy) and had a positive oro/nasopharyngeal swab reverse‐transcription polymerase chain reaction result. The highest sensitivity with a very good specificity performance was reached at a cutoff value of 10.0 AU/mL for IgM and of 7.1 for IgG antibodies, hence near to the manufacturer's cutoff values of 10 AU/mL for both isotypes. The receiver operating characteristic curves showed area under the curve values of 0.918 and 0.980 for anti‐SARS CoV‐2 antibodies IgM and IgG, respectively. iFlash1800 CLIA analyzer has shown highly accurate results for the anti‐SARS‐CoV‐2 antibodies profile and can be considered an excellent tool for COVID‐19 diagnostics.

the need to accelerate progress in diagnostics, serological tests have been developed. More than 200 different assays have been proposed so far but almost all have poor regulatory status and lack clinical and analytical performance review. 8 In fact the speed with which they are released in the market and the versatility of immunoassays such as source of antigen and secondary antibody conjugate, make them poorly evaluated tests. Given that during the outbreak test validation is not a priority and given that nonlaboratory specialists are allowed to handle these tests because of limited staff resources has meant that unregulated testing has spread widely. In particular, since rapid tests do not require any instruments or laboratory personnel they could be set up anywhere and at any time, especially in developing nations with limited healthcare resources and in remote settings. The more relaxed rules of the FDA's "Policy for Diagnostic Tests for Coronavirus Disease-2019 during the Public Health Emergency" issued on 16 March 2020, 9 has allowed the market easier access to these tests as well as easier and faster diagnostics, but the lack of control in the production process is also dangerous making these tests potentially less reliable. Along with chromatographic rapid immunoassays as qualitative tests, 10 quantitative antibodies detection tests, such as enzyme-linked immunosorbent assay and chemiluminescence immunoassay (CLIA), have spread often by fully automated analyzers.

These technologies characterized by high-throughput and low complexity have helped us to use serological testing more accurately during both antibody development and monitoring the different phases of the disease. Indeed, being able to receive information about the antibody concentration and time kinetics of humoral response is very important for diagnostic, prognostic, and therapeutic applications. 11 The aim of the this study was to assess the diagnostic performance of a novel fully automated CLIA for the quantitative detection of anti-SARS-CoV-2 IgM and IgG antibodies. [12] [13] [14] [15] However, it is unclear which antibodies are optimally effective in the scenario of COVID-19 and which of them are neutralizing. There is also uncertainty as to which antibody isotype (IgM, IgG or IgA) (single or combined) is the best choice in these different contexts. 16 As with most existing studies on the diagnostic performance of the SARS-CoV-2 antibodies, our preliminary data showed that most COVID-19 patients have both IgM and IgG, and only few of them have isolated IgG or IgM antibodies. On the one hand, in reference to IgM and IgG combination, the overall sensitivity of 75% may reflect that some patients may not yet develop antibodies or will never develop (the length of time from the symptoms onset to serological test ranged from 8 to 17 days); on the other hand, the 100% specificity performance of IgG antibodies makes them an appropriate test for the different immunization protocols. With regard to IgM false positive results, it's important to underline that we designed a disease control group made up of (a) donors from last winter when other coronaviruses were active who had all negative results;

(b) autoimmune and infectious diseases dating back at least 1 year in which we found four reactive sera. This means that we had no cross reaction with other coronaviruses but two CMV infections and two rheumatic diseases interfered with the test, even if with a low titer.

This data can be added to the known issues concerning IgM by rapid tests such as the lack of specificity together with the low sensitivity 

Our experience highlights the importance of a CLIA method, not only to overcome the problems of the subjective reading of the band (especially weak) in the rapid tests, but for the wide range of potentials inherent to a quantitative method, such as assisting with diagnosis and evaluating the disease through antibodies profiles.

Furthermore, selection of IgG antibodies at high level concentrations may be helpful in developing vaccines and treating SARS-CoV-2 by convalescent plasma therapy. Abbreviations: CLIA, chemiluminescence immunoassay; LR + , positive likelihood ratio; LR−, negative likelihood ratio; NPV, negative predictive value; OR, odds ratio; PPV, positive predictive value.

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