Browse
Submit Data
Databases
API
Help

Dataset Information

13 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Monitoring Neutralization Property Change of Evolving Hantaan and Seoul Viruses with a Novel Pseudovirus-Based Assay.

ABSTRACT: The Hantaan virus (HTNV) and Seoul virus (SEOV) mutants have accumulated over time. It is important to determine whether their neutralizing epitopes have evolved, thereby making the current vaccine powerless. However, it is impossible to determine by using traditional plaque reduction neutralization test (PRNT), because it requires large numbers of live mutant strains. Pseudovirus-based neutralization assays (PBNA) were developed by employing vesicular stomatitis virus (VSV) backbone incorporated with HTNV or SEOV glycoproteins (VSVΔG^*-HTNVG or VSVΔG^*-SEOVG). 56 and 51 single amino acid substitutions of glycoprotein (GP) in HTNV and SEOV were selected and introduced into the reference plasmid. Then the mutant pseudoviruses were generated and tested by PBNA. The PBNA results were highly correlated with PRNT ones with R² being 0.91 for VSVΔG^*-HTNVG and 0.82 for VSVΔG^*-SEOVG. 53 HTNV mutant pseudoviruses and 46 SEOV mutants were successfully generated. Importantly, by using PBNA, we found that HTNV or SEOV immunized antisera could neutralize all the corresponding 53 HTNV mutants or the 46 SEOV mutants respectively. The novel PBNA enables us to closely monitor the effectiveness of vaccines against large numbers of evolving HTNV and SEOV. And the current vaccine remains to be effective for the naturally occurring mutants.

SUBMITTER: Ning T

PROVIDER: S-EPMC7973352 | biostudies-literature |

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Similar Datasets

A Simplified SARS-CoV-2 Pseudovirus Neutralization Assay.

Project description:COVID-19 is an ongoing pandemic caused by the highly infectious coronavirus SARS-CoV-2 that is engaging worldwide scientific research to find a timely and effective eradication strategy. Great efforts have been put into anti-COVID-19 vaccine generation in an effort to protect the world population and block SARS-CoV-2 spread. To validate the protective efficacy of the vaccination campaign and effectively control the pandemic, it is necessary to quantify the induction of neutralizing antibodies by vaccination, as they have been established to be a correlate of protection. In this work, a SARS-CoV-2 pseudovirus neutralization assay, based on a replication-incompetent lentivirus expressing an adapted form of CoV-2 S protein and an ACE2/TMPRSS2 stably expressing cell line, has been minimized in terms of protocol steps without loss of accuracy. The goal of the present simplified neutralization system is to improve SARS-CoV-2 vaccination campaign by means of an easy and accessible approach to be performed in any medical laboratory, maintaining the sensitivity and quantitative reliability of classical serum neutralization assays. Further, this assay can be easily adapted to different coronavirus variants by simply modifying the pseudotyping vector.

| S-EPMC8071407 | biostudies-literature

An HFman Probe-Based Multiplex Reverse Transcription Loop-Mediated Isothermal Amplification Assay for Simultaneous Detection of Hantaan and Seoul Viruses.

Project description:Hantaviruses are zoonotic pathogens that are widely distributed worldwide. Hantaan virus (HTNV) and Seoul virus (SEOV) are two most common hantaviruses that infect humans and cause hemorrhagic fever with renal syndrome (HFRS). Rapid and sensitive detection of HTNV and SEOV are crucial for surveillance, clinical treatment and management of HFRS. This study aimed to develop a rapid HFman probe-based mulstiplex reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay to simultaneously detect HTNV and SEOV. A novel multiplex RT-LAMP assay was developed, and 46 serum samples obtained from clinically suspected patients were used for evaluation. The novel RT-LAMP assay can detect as low as 3 copies/reaction of hantaviruses with a detection limit of 41 and 73 copies per reaction for HTNV and SEOV, respectively. A clinical evaluation showed that the consistencies of the multiplex RT-LAMP with RT-qPCR assay were 100% and 97.8% for HTNV and SEOV, respectively. In view of the high prevalence of HTNV and SEOV in rural areas with high rodent density, a colorimetric visual determination method was also developed for point-of-care testing (POCT) for the diagnosis of the two viruses. The novel multiplex RT-LAMP assay is a sensitive, specific, and efficient method for simultaneously detecting HTNV and SEOV.

| S-EPMC9406646 | biostudies-literature

Evaluation of a Pseudovirus Neutralization Assay for SARS-CoV-2 and Correlation with Live Virus-Based Micro Neutralization Assay.

Project description:The unusual cases of pneumonia outbreak were reported from Wuhan city in late December 2019. Serological testing provides a powerful tool for the identification of prior infection and for epidemiological studies. Pseudotype virus neutralization assays are widely used for many viruses and applications in the fields of serology. The accuracy of pseudotype neutralizing assay allows for its use in low biosafety lab and provides a safe and effective alternative to the use of wild-type viruses. In this study, we evaluated the performance of this assay compared to the standard microneutralization assay as a reference. The lentiviral pseudotype particles were generated harboring the Spike gene of SARS-CoV-2. The generated pseudotype particles assay was used to evaluate the activity of neutralizing antibodies in 300 human serum samples from a COVID-19 sero-epidemiological study. Testing of these samples resulted in 55 positive samples and 245 negative samples by pseudotype viral particles assay while microneutralization assay resulted in 64 positive and 236 negative by MN assay. Compared to the MN, the pseudotyped viral particles assay showed a sensitivity of 85.94% and a specificity of 100%. Based on the data generated from this study, the pseudotype-based neutralization assay showed a reliable performance for the detection of neutralizing antibodies against SARS-CoV-2 and can be used safely and efficiently as a diagnostic tool in a biosafety level 2 laboratory.

| S-EPMC8226551 | biostudies-literature

Safe pseudovirus-based assay for neutralization antibodies against influenza A(H7N9) virus.

Project description:Serologic studies are urgently needed to assist in understanding an outbreak of influenza A(H7N9) virus. However, a biosafety level 3 laboratory is required for conventional serologic assays with live lethal virus. We describe a safe pseudovirus-based neutralization assay with preliminary assessment using subtype H7N9-infected samples and controls.

| S-EPMC3810762 | biostudies-literature

Developing Pseudovirus-Based Neutralization Assay against Omicron-Included SARS-CoV-2 Variants.

Project description:The global spread of SARS-CoV-2 and its variants poses a serious threat to human health worldwide. Recently, the emergence of Omicron has presented a new challenge to the prevention and control of the COVID-19 pandemic. A convenient and reliable in vitro neutralization assay is an important method for validating the efficiency of antibodies, vaccines, and other potential drugs. Here, we established an effective assay based on a pseudovirus carrying a full-length spike (S) protein of SARS-CoV-2 variants in the HIV-1 backbone, with a luciferase reporter gene inserted into the non-replicate pseudovirus genome. The key parameters for packaging the pseudovirus were optimized, including the ratio of the S protein expression plasmids to the HIV backbone plasmids and the collection time for the Alpha, Beta, Gamma, Kappa, and Omicron pseudovirus particles. The pseudovirus neutralization assay was validated using several approved or developed monoclonal antibodies, underscoring that Omicron can escape some neutralizing antibodies, such as REGN10987 and REGN10933, while S309 and ADG-2 still function with reduced neutralization capability. The neutralizing capacity of convalescent plasma from COVID-19 convalescent patients in Wuhan was tested against these pseudoviruses, revealing the immune evasion of Omicron. Our work established a practical pseudovirus-based neutralization assay for SARS-CoV-2 variants, which can be conducted safely under biosafety level-2 (BSL-2) conditions, and this assay will be a promising tool for studying and characterizing vaccines and therapeutic candidates against Omicron-included SARS-CoV-2 variants.

| S-EPMC9231177 | biostudies-literature

Establishment and validation of a pseudovirus neutralization assay for SARS-CoV-2.

Project description:Pseudoviruses are useful virological tools because of their safety and versatility, especially for emerging and re-emerging viruses. Due to its high pathogenicity and infectivity and the lack of effective vaccines and therapeutics, live SARS-CoV-2 has to be handled under biosafety level 3 conditions, which has hindered the development of vaccines and therapeutics. Based on a VSV pseudovirus production system, a pseudovirus-based neutralization assay has been developed for evaluating neutralizing antibodies against SARS-CoV-2 in biosafety level 2 facilities. The key parameters for this assay were optimized, including cell types, cell numbers, virus inoculum. When tested against the SARS-CoV-2 pseudovirus, SARS-CoV-2 convalescent patient sera showed high neutralizing potency, which underscore its potential as therapeutics. The limit of detection for this assay was determined as 22.1 and 43.2 for human and mouse serum samples respectively using a panel of 120 negative samples. The cutoff values were set as 30 and 50 for human and mouse serum samples, respectively. This assay showed relatively low coefficient of variations with 15.9% and 16.2% for the intra- and inter-assay analyses respectively. Taken together, we established a robust pseudovirus-based neutralization assay for SARS-CoV-2 and are glad to share pseudoviruses and related protocols with the developers of vaccines or therapeutics to fight against this lethal virus.

| S-EPMC7144318 | biostudies-literature

Establishment of a neutralization assay for Nipah virus using a high-titer pseudovirus system.

Project description:ObjectiveTo construct a high-titer Nipah pseudovirus packaging system using the HIV lentivirus backbone vector and establish a safe neutralization assay for Nipah pseudovirus in biosafety level 2 facilities.MethodsNipah virus (NiV) fusion protein (F) and glycoprotein (G) recombinant expression plasmids, psPAX2, and pLenti CMV Puro LUC (w168-1) were transiently transfected into 293T cells for 72 h for the generation of a NiV pseudovirus. The neutralization ability of Nipah virus F and G protein antibodies was assessed using the pseudovirus.ResultsA NiV pseudovirus was constructed using 293T cells. The ideal mass ratio of plasmid psPAX2: w168-1: F: G for transfection was determined to be 4:4:1:1. The specificity of recombinant F and G protein expression was indicated by indirect immunofluorescence and western blotting. The pseudovirus particles showed obvious spikes under a transmission electron microscope. The NiV pseudovirus titer was 4.73 × 105 median tissue culture infective dose per mL, and the pseudovirus could be effectively neutralized by polyclonal antibodies specifically targeting the F and G proteins respectively.ConclusionsA NiV pseudovirus was successfully generated using HIV vector systems, and was used as a platform for a safe and reliable pseudovirus-based neutralizing assay that can be performed in biosafety level 2 facilities.

| S-EPMC9860241 | biostudies-literature

Establishment of a pseudovirus neutralization assay based on SARS-CoV-2 S protein incorporated into lentiviral particles.

Project description:The coronavirus disease 2019 (COVID-19) is still causing a wide range of infections and deaths due to the high variability of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Therefore, it is necessary to establish a reliable and convenient pseudovirus-based neutralization assay to develop drug targeted variants of SARS-CoV-2. Based on the HIV-1 backbone, we generated a high titer luciferase (Luc)-expressing pseudovirus packaging system. Three dominant S mutant substitution pseudovirus were also established and identified compared to wide type in hACE2-overexpressing HEK-293T cells (293T-ACE2 cells). Compared to serine protease inhibitor camostat mesylate, the cysteine protease inhibitor E-64d could significantly block all SARS-CoV-2 mutant S pseudovirus infection in 293T-ACE2 cells. Furthermore, the neutralization ability of two antibodies targeted receptor-binding domain (RBD) of SARS-CoV-2 spike protein (S) was evaluated, which showed different inhibition dose-effect curves among four types of S pseudovirus. Overall, we developed a pseudovirus-based neutralization assay for SARS-CoV-2, which would be readily adapted to SARS-CoV-2 variants for evaluating antibodies.

| S-EPMC8721934 | biostudies-literature

SARS-CoV-2 Antibody Neutralization Assay Platforms Based on Epitopes Sources: Live Virus, Pseudovirus, and Recombinant S Glycoprotein RBD.

Project description:The high virulent severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) virus that emerged in China at the end of 2019 has generated novel coronavirus disease, coronavirus disease 2019 (COVID-19), causing a pandemic worldwide. Every country has made great efforts to struggle against SARS-CoV-2 infection, including massive vaccination, immunological patients' surveillance, and the utilization of convalescence plasma for COVID-19 therapy. These efforts are associated with the attempts to increase the titers of SARS-CoV-2 neutralizing Abs (nAbs) generated either after infection or vaccination that represent the body's immune status. As there is no standard therapy for COVID-19 yet, virus eradication will mainly depend on these nAbs contents in the body. Therefore, serological nAbs neutralization assays become a requirement for researchers and clinicians to measure nAbs titers. Different platforms have been developed to evaluate nAbs titers utilizing various epitopes sources, including neutralization assays based on the live virus, pseudovirus, and neutralization assays utilizing recombinant SARS-CoV-2 S glycoprotein receptor binding site, receptor-binding domain. As a standard neutralization assay, the plaque reduction neutralization test (PRNT) requires isolation and propagation of live pathogenic SARS-CoV-2 virus conducted in a BSL-3 containment. Hence, other surrogate neutralization assays relevant to the PRNT play important alternatives that offer better safety besides facilitating high throughput analyses. This review discusses the current neutralization assay platforms used to evaluate nAbs, their techniques, advantages, and limitations.

| S-EPMC8733193 | biostudies-literature

Simultaneous rapid detection of Hantaan virus and Seoul virus using RT-LAMP in rats.

Project description:BackgroundHemorrhagic fever with renal syndrome is in most cases caused by the Hantaan virus (HTNV) and Seoul virus (SEOV). To develop and apply reverse transcription loop-mediated isothermal amplification (RT-LAMP) to detect HTNV and SEOV simultaneously, which was faster, more cost effective, and easier to perform as the target gene amplified rapidly. In this article an assay based on LAMP is demonstrated, which only employs such apparatus as a water bath or a heat block.MethodsA chromogenic method using the calcein/Mn2+ complex and real-time turbidity monitoring method were used to assess reaction progress of the reaction, and the specificity of the RT-LAMP-based assay was assessed by detecting cDNAs/cRNAs generated from Coxsackievirus A16, Influenza virus, lymphocytic choriomeningitis virus, mouse poxvirus, rotavirus, mouse hepatitis virus. In addition, 23 clinical specimens were used to determine the agreement between the RT-LAMP assay with reverse transcriptase polymerase chain reaction (RT-PCR) and immunofluorescence (IFT) method.ResultsThe detection limit of RT-LAMP to HNTV and SEOV was as low as 10 copies/μL with optimized reaction conditions, which was much more sensitive than the RT-PCR method (100-1,000 copies/μL). At the same time, the detection results of 23 clinical specimens have also illustrated the agreement between this the RT-LAMP assay with RT-PCR and IFT.DiscussionThis RT-LAMP assay could be used to perform simultaneous and rapid detection of HTNV and SEOV to the clinical specimens.

| S-EPMC6329334 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data