Neutralization assay with SARS-CoV-1 and SARS-CoV-2 spike pseudotyped murine leukemia virions

Abstract

Background Virus neutralization by antibodies is an important prognostic factor in many viral diseases. To easily and rapidly measure titers of neutralizing antibodies in serum or plasma, we developed pseudovirion particles composed of the spike glycoprotein of SARS-CoV-2 incorporated onto murine leukemia virus capsids and a modified minimal murine leukemia virus genome encoding firefly luciferase. This assay design is intended for use in laboratories with biocontainment level 2 and therefore circumvents the need for the biocontainment level 3 that would be required for replication-competent SARS-CoV-2 virus. To validate the pseudovirion assay, we set up comparisons with other available antibody tests including those from Abbott, Euroimmun and Siemens, using archived, known samples. Results 11 out of 12 SARS-CoV-2-infected patient serum samples showed neutralizing activity against SARS-CoV-2-spike pseudotyped MLV viruses, with neutralizing titers-50 (NT50) that ranged from 1:25 to 1:1,417. Five historical samples from patients hospitalized for severe influenza infection in 2016 tested negative in the neutralization assay (NT50 < 25). Three serum samples with high neutralizing activity against SARS-CoV-2/MLV pseudoviruses showed no detectable neutralizing activity (NT50 < 25) against SARS-CoV-1/MLV pseudovirions. We also compared the semiquantitative Siemens SARS-CoV-2 IgG test, which measures binding of IgG to recombinantly expressed receptor binding domain of SARS-CoV-2 spike glycoprotein with the neutralization titers obtained in the pseudovirion assay and the results show high concordance between the two tests (R2 = 0.9344). Conclusions SARS-CoV-2 spike/MLV pseudovirions provide a practical means of assessing neutralizing activity of antibodies in serum or plasma from infected patients under laboratory conditions consistent with biocontainment level 2. This assay offers promise also in evaluating immunogenicity of spike glycoprotein-based candidate vaccines in the near future.

Superiority of MALDI-TOF Mass Spectrometry over Real-Time PCR for SARS-CoV-2 RNA Detection

“…Our data analysis is concordant with other studies reporting false-negative results from RT-PCR tests ranging up to 30% [ 4 ]. Several studies have already showed that the diagnostic efficacy between RT-PCR test kits for SARS-CoV-2 nucleic acid detection may differ significantly depending on the probes and primers used [ 12 , 13 ].…”

“…According to the WHO and Chinese Center for Disease Control and Prevention (CDC), real-time PCR (RT-PCR) is considered the ‘gold standard’ method for detection of SARS-CoV-2 infection [ 3 ]. Nevertheless, studies have shown that RT-PCR is an error-prone method showing false-negative results rates from respiratory samples for SARS-CoV-2 ranging from 1% to 30% [ 4 ]. These diagnostic errors are related to a number of preanalytical and analytical factors, including the source of the sample, delays or inadequate storage conditions before arrival to the laboratory, possible errors of RNA extractions, as well as the different quality and sensitivity of detection kits [ 5 ].…”

“…These diagnostic errors are related to a number of preanalytical and analytical factors, including the source of the sample, delays or inadequate storage conditions before arrival to the laboratory, possible errors of RNA extractions, as well as the different quality and sensitivity of detection kits [ 5 ]. Additionally, insufficient viral load, the incubation period of the disease, and the presence of mutations that allow virus to escape from the detection may also lead to the risk of obtaining false-negative results [ 4 ]. Therefore, the improvement of the COVID-19 diagnosis is urgently needed for obtaining true negative results, allowing one to exit quarantine and avoid virus transmission and recurrence.…”

Profile of Circulatory Cytokines and Chemokines in Human Coronaviruses: A Systematic Review and Meta-Analysis

“…It is worth noting that CXCL10/IP10 is absent in healthy individuals. However, it is detected in asymptomatic patients and is directly linked to disease severity ( 96 , 97 ), suggesting that early detection of chemokines can help control new outbreaks ( 98 ). Several clinical investigations involving autopsies have also provided evidence that thrombosis is an important consequence of hCoV diseases ( 99 ).…”

Viral transport media for COVID-19 testing

“…COVID-19 test kits employed by our group supported the propagation of local Columbus, OH strains of SARS-CoV-2 from testing samples, confirming that our VTM formulation maintained infectious virions. In laboratory experiments, others have shown that live SARS-CoV-2 survives in similar VTM for at least 72 h at room temperature, and viral RNA can be detected by RT-PCR as long as 28 days post collection [ 6 , 7 ]. In agreement with this, OSUWMC has contributed numerous full genome sequences of SARS-CoV-2 obtained from our testing kit samples, and identified a Midwestern variant of concern [8] .…”