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Evaluation of Two Chemiluminescent and Three ELISA Immunoassays for the Detection of SARS-CoV-2 IgG Antibodies: Implications for Disease Diagnosis and Patients’ Management

ABSTRACT: The estimation of anti-SARS-CoV-2 IgG antibodies is possibly the best approach to accurately establish the number of infected individuals and the seroprevalence of COVID-19 within a population. Thus, several commercial immunoassays have recently been developed. The purpose of our study was to assess the performance of five commonly used immunoassays in Greece (3 ELISA, namely Euroimmun SARS-CoV-2, GA GENERIC SARS-CoV-2 and Vircell COVID-19; and 2 chemiluminescent, namely ABBOTT SARS-CoV-2 and ROCHE Elecsys Anti-SARS-CoV-2 test) for the detection of anti-SARS-CoV-2 IgG antibodies. Sera specimens derived from 168 individuals were utilized to assess the specificity and sensitivity score of each assay. Among them, we included 99 COVID-19 patients (29 asymptomatic, 36 with symptom onset 4 to 14 days before serum sampling, and 34 with symptom initiation ? 15 days ago), and 69 volunteers with sera specimens collected prior to the SARS-CoV-2 outbreak and maintained at ?80°C. We demonstrated that chemiluminescent immunoassays exhibit a significantly higher specificity score but a lower sensitivity, compared to ELISA immunoassays. Moreover, immunoassays detecting IgG antibodies against SARS-CoV-2 N protein instead of S protein alone are more reliable, considering both specificity and sensitivity scores. Interestingly, all asymptomatic patients displayed anti-SARS-CoV-2 IgG antibodies, confirmed by at least two immunoassays. We suggest that chemiluminescent assays could be used as screening methods for the detection of anti-SARS-CoV-2 antibodies to evaluate the possible prevalence of disease in the general population, while ELISA assays would be more reliable to evaluate, and follow-up confirmed COVID-19 patients.

SUBMITTER: Speletas M

PROVIDER: S-EPMC7785794 | biostudies-literature | 2020 Jan

REPOSITORIES: biostudies-literature

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Project description:BackgroundA novel porcine deltacoronavirus (PDCoV), also known as porcine coronavirus HKU15, was reported in China in 2012 and identified in the U.S. in early 2014. Since then, PDCoV has been identified in a number of U.S. states and linked with clinical disease including acute diarrhea and vomiting in the absence of other identifiable pathogens. Since PDCoV was just recently linked with clinical disease, few specific antibody-based reagents were available to assist in diagnosis of PDCoV and limited serological capabilities were available to detect an antibody response to this virus. Therefore, the overall objective of this project was to develop and validate selected diagnostic reagents and assays for PDCoV antigen and antibody detection.ResultsThe nucleoprotein of PDCoV was expressed as a recombinant protein and purified for use as an antigen to immunize mice for polyclonal, hyperimmune sera and monoclonal antibody (mAb) production. The resulting mAbs were evaluated for use in fluorescent antibody staining methods to detect PDCoV infected cells following virus isolation attempts and for immunohistochemistry staining of intestinal tissues of infected pigs. The same antigen was used to develop serological tests to detect the antibody response to PDCoV in pigs following infection. Serum samples from swine herds with recent documentation of PDCoV infection and samples from expected naïve herds were used for initial assay optimization. The tests were optimized in a checkerboard fashion to reduce signal to noise ratios using samples of known status. Statistical analysis was performed to establish assay cutoff values and assess diagnostic sensitivities and specificities. At least 629 known negative serum samples and 311 known positive samples were evaluated for each assay. The enzyme linked immunosorbent assay (ELISA) showed diagnostic sensitivity (DSe) of 96.1% and diagnostic specificity (DSp) of 96.2%. The fluorescent microsphere immunoassay (FMIA) showed a DSe of 95.8% and DSp of 98.1%. Both ELISA and FMIA detected seroconversion of challenged pigs between 8-14 days post-infection (DPI). An indirect fluorescent antibody (IFA) test was also developed using cell culture adapted PDCoV for comparative purposes.ConclusionThese new, specific reagents and serological assays will allow for improved diagnosis of PDCoV. Since many aspects of PDCoV infection and transmission are still not fully understood, the reagents and assays developed in this project should provide valuable tools to help understand this disease and to aid in the control and surveillance of porcine deltacoronavirus outbreaks.

Project description:The severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) pandemic has adversely affected humankind and caused millions of deaths globally since January 2020. Robust and quick serological tests such as antibody detection assays for SARS-CoV-2 provide relevant information and aid in the process of vaccine development and diagnostics, as well as in sero-epidemiological monitoring of antibody response to the virus. The receptor-binding domain (RBD) of spike and nucleocapsid protein are specific targets for detecting SARS-CoV-2 antibodies. Here, we present the development of a stable spike (S) and nucleocapsid (N) protein-based ELISA antibody detection test "CoroSuchak," with 99% sensitivity, 98% specificity, cost-effective, and detection in a minimum time for serodiagnosis and mass screening of the population for antibodies against SARS-CoV-2. Blood samples were analyzed from 374 SARS-CoV-2 reverse transcription-polymerase chain reaction (RT-PCR) positive, 772 negative and asymptomatic, and 874 random groups of subjects. We found that the antibody titer was significantly higher (p < 0.0001) in infected and vaccinated group compared to the only vaccinated and only infected group. Using enzyme-linked immunosorbent assay (ELISA), we detected SARS-CoV-2 immunoglobulin G (IgG) antibodies in 118/123 (96%) infected individuals, 570/653 (87%) non-infected but vaccinated individuals, 231/237 (97%) individuals who were both infected and vaccinated, and 499/874 (57%) from randomly selected individuals from the first and second waves of the pandemic. Similarly in the third wave, 14/14 (100%) infected and 16/20 (80%) RT-PCR-negative but symptomatic subjects were detected. Thus, the highly sensitive and specific in-house developed ELISA antibody detection kit "CoroSuchak" is extremely useful to determine the seroprevalence of SARS-CoV-2 antibodies in the coronavirus-exposed population. KEY POINTS: •Indigenous kit using a combination of spike and nucleocapsid proteins and peptide sequences. •High sensitivity and specificity to detect variants. •Highly sensitive for mass screening.

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Evaluation of Two Chemiluminescent and Three ELISA Immunoassays for the Detection of SARS-CoV-2 IgG Antibodies: Implications for Disease Diagnosis and Patients’ Management

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