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Single-reaction multi-antigen serological test for comprehensive evaluation of SARS-CoV-2 patients by flow cytometry.

ABSTRACT: Here, we describe a new, simple, highly multiplexed serological test that generates a more complete picture of seroconversion than single antigen-based assays. Flow cytometry is used to detect multiple Ig isotypes binding to four SARS-CoV-2 antigens: the Spike glycoprotein, its RBD fragment (the main target for neutralizing antibodies), the nucleocapsid protein, and the main cysteine-like protease in a single reaction. Until now, most diagnostic serological tests measured antibodies to only one antigen and in some laboratory-confirmed patients no SARS-CoV-2-specific antibodies could be detected. Our data reveal that while most patients respond against all the viral antigens tested, others show a marked bias to make antibodies against either proteins exposed on the viral particle or those released after cellular infection. With this assay, it was possible to discriminate between patients and healthy controls with 100% confidence. Analysing the response of multiple Ig isotypes to the four antigens in combination may also help to establish a correlation with the severity degree of disease. A more detailed description of the immune responses of different patients to SARS-CoV-2 virus might provide insight into the wide array of clinical presentations of COVID-19.

SUBMITTER: Caceres-Martell Y

PROVIDER: S-EPMC8420214 | biostudies-literature | 2021 Nov

REPOSITORIES: biostudies-literature

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Single-reaction multi-antigen serological test for comprehensive evaluation of SARS-CoV-2 patients by flow cytometry.

Cáceres-Martell Yaiza Y Fernández-Soto Daniel D Campos-Silva Carmen C García-Cuesta Eva M EM Casasnovas Jose M JM Navas-Herrera David D Beneítez-Martínez Alexandra A Martínez-Fleta Pedro P Alfranca Arantzazu A Sánchez-Madrid Francisco F Escudero-López Gabriela G Vilches Carlos C Jara-Acevedo Ricardo R Reyburn Hugh T HT Rodríguez-Frade José M JM Valés-Gómez Mar M

European journal of immunology 20210819 11

Here, we describe a new, simple, highly multiplexed serological test that generates a more complete picture of seroconversion than single antigen-based assays. Flow cytometry is used to detect multiple Ig isotypes binding to four SARS-CoV-2 antigens: the Spike glycoprotein, its RBD fragment (the main target for neutralizing antibodies), the nucleocapsid protein, and the main cysteine-like protease in a single reaction. Until now, most diagnostic serological tests measured antibodies to only one ...[more]

PMID: 34358329

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Project description:The onset of the coronavirus disease 2019 (COVID-19) pandemic resulted in hundreds of in vitro diagnostic devices (IVDs) coming to market, facilitated by regulatory authorities allowing "emergency use" without a comprehensive evaluation of performance. The World Health Organization (WHO) released target product profiles (TPPs) specifying acceptable performance characteristics for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) assay devices. We evaluated 26 rapid diagnostic tests and 9 enzyme immunoassays (EIAs) for anti-SARS-CoV-2, suitable for use in low- and middle-income countries (LMICs), against these TPPs and other performance characteristics. The sensitivity and specificity ranged from 60.1 to 100% and 56.0 to 100%, respectively. Five of 35 test kits reported no false reactivity for 55 samples with potentially cross-reacting substances. Six test kits reported no false reactivity for 35 samples containing interfering substances, and only one test reported no false reactivity with samples positive for other coronaviruses (not SARS-CoV-2). This study demonstrates that a comprehensive evaluation of the performance of test kits against defined specifications is essential for the selection of test kits, especially in a pandemic setting. IMPORTANCE The markets have been flooded with hundreds of SARS-CoV-2 serology tests, and although there are many published reports on their performance, comparative reports are far fewer and tend to be limited to only a few tests. In this report, we comparatively assessed 35 rapid diagnostic tests or microtiter plate enzyme immunoassays (EIAs) using a large set of samples from individuals with a history of mild to moderate COVID-19, commensurate with the target population for serosurveillance, which included serum samples from individuals previously infected, at undetermined time periods, with other seasonal human coronaviruses, Middle East respiratory syndrome coronavirus (MERS-CoV), and SARS-CoV-1. The significant heterogeneity in their performances, with only a few tests meeting WHO target product profile performance requirements, highlights the importance of independent comparative assessments to inform the use and procurement of these tests for both diagnostics and epidemiological investigations.

Project description:BackgroundViral diversity presents an ongoing challenge for diagnostic tests, which need to accurately detect all circulating variants. The Abbott Global Surveillance program monitors severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) variants and their impact on diagnostic test performance.ObjectivesTo evaluate the capacity of Abbott molecular, antigen, and serologic assays to detect circulating SARS-CoV-2 variants, including all current variants of concern (VOC): B.1.1.7 (alpha), B.1.351 (beta), P.1 (gamma) and B.1.617.2 (delta).Study designDilutions of variant virus cultures (B.1.1.7, B.1.351, B.1.429, B.1.526.1, B.1.526.2, B.1.617.1, B.1.617.2, P.1, R.1 and control isolate WA1) and a panel of N = 248 clinical samples from patients with sequence confirmed variant infections (B.1.1.7, B.1.351, B.1.427, B.1.429, B.1.526, B.1.526.1, B.1.526.2, P.1, P.2, R.1) were evaluated on at least one assay: Abbott ID NOW COVID-19, m2000 RealTime SARS-CoV-2, Alinity m SARS-CoV-2, and Alinity m Resp-4-Plex molecular assays; the BinaxNOW COVID-19 Ag Card and Panbio COVID-19 Ag Rapid Test Device; and the ARCHITECT/Alinity i SARS-CoV-2 IgG and AdviseDx IgM assays, Panbio COVID-19 IgG assay, and ARCHITECT/Alinity i AdviseDx SARS-CoV-2 IgG II assay.ResultsConsistent with in silico predictions, each molecular and antigen assay detected VOC virus cultures with equivalent sensitivity to the WA1 control strain. Notably, 100% of all tested variant patient specimens were detected by molecular assays (N = 197 m2000, N = 88 Alinity m, N = 99 ID NOW), and lateral flow assays had a sensitivity of >94% for specimens with genome equivalents (GE) per device above 4 log (85/88, Panbio; 54/57 Binax). Furthermore, Abbott antibody assays detected IgG and IgM in 94-100% of sera from immune competent B.1.1.7 patients 15-26 days after symptom onset.ConclusionsThese data confirm variant detection for 11 SARS-CoV-2 assays, which is consistent with each assay target region being highly conserved. Importantly, alpha, beta, gamma, and delta VOCs were detected by molecular and antigen assays, indicating that these tests may be suitable for widescale use where VOCs predominate.

Dataset Information

Single-reaction multi-antigen serological test for comprehensive evaluation of SARS-CoV-2 patients by flow cytometry.

Publications

Single-reaction multi-antigen serological test for comprehensive evaluation of SARS-CoV-2 patients by flow cytometry.

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