Ontology highlight
ABSTRACT:
PROVIDER: PRJNA1037295 | ENA |
REPOSITORIES: ENA
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Project description:Qiagen RT-PCR array Dimethylamin validation experiment
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Project description:There is an urgent need for robust and high-throughput methods for SARS-CoV-2 detection in suspected pa-tient samples to facilitate disease management, surveillance, and control. Although nucleic acid detection methods such as RT-PCR are the gold standard, during the current pandemic the deployment of RT-PCR tests has been extremely slow, and key reagents such as PCR primers, and RNA extraction kits are at critical shortages. Rapid point-of-care viral antigen detec-tion methods have been previously employed for the diagnosis of respiratory viruses such as influenza and respiratory syn-cytial viruses. Therefore, the direct detection of SARS-CoV-2 viral antigens in patient samples could also be used for diagno-sis of active infection and alternative methodologies for specific and sensitive viral protein detection should be explored. Targeted mass spectrometry techniques have enabled the identification and quantitation of a defined subset of pro-teins/peptides at single amino acid resolution with attomole level sensitivity and high reproducibility. Herein we report a tar-geted mass spectrometry assay for the detection of SARS- CoV-2 spike protein and nucleoprotein in a relevant biologi-cal matrix. Recombinant full-length spike protein and nucleoprotein were digested and prototypic peptides were selected for parallel reaction monitoring (PRM) quantitation using a high resolution Orbitrap instrument. A spectral library, which con-tained 7 proteotypic peptides (4 from spike protein and 3 from nucleoprotein) and the top 3 to 4 transitionsMS2 spectra, was generated and evaluated. From the original spectral library, we selected 2 best performing peptides for the final PRM assay. The assay was evaluated using mock test samples containing inactivated SARS-CoV-2 virions, added to in-vitro de-rived mucus. The PRM assay provided a limit of detection (LOD) of ~200 attomoles and a limit of quantitation (LOQ) of ~ 390 attomoles. Extrapolating from the test samples, the projected titer of virus particles necessary for detection of SARS-CoV-2 spike and nucleoprotein detection was approximately 2E5 viral particles/mL, making it an attractive alternative to RT-PCR assays. Potentially mass spectrometry-based methods for viral antigen detection may deliver higher throughput and could serve as a complementary diagnostic tool to RT-PCR. Furthermore, this assay could be used to evaluate the pres-ence of SARS-CoV-2 in archived or recently collected biological fluids, in-vitro derived research materials, and wastewater samples
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Project description:Serum miRNA profiling by quantitative RT PCR [Validation]
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Project description:Detection of SARS-CoV-2 using RT–PCR and other advanced methods can achieve high accuracy. However, their application is limited in countries that lack sufficient resources to handle large-scale testing during the COVID-19 pandemic. Here, we describe a method to detect SARS-CoV-2 in nasal swabs using matrix-assisted laser desorption/ionization mass spectrometry (MALDI-MS) and machine learning analysis. This approach uses equipment and expertise commonly found in clinical laboratories in developing countries. We obtained mass spectra from a total of 362 samples (211 SARS-CoV-2-positive and 151 negative by RT–PCR) without prior sample preparation from three different laboratories. We tested two feature selection methods and six machine learning approaches to identify the top performing analysis approaches and determine the accuracy of SARS-CoV-2 detection. The support vector machine model provided the highest accuracy (93.9%), with 7% false positives and 5% false negatives. Our results suggest that MALDI-MS and machine learning analysis can be used to reliably detect SARS-CoV-2 in nasal swab samples.
2020-09-11 | PXD021388 | Pride
Project description:SARS-CoV-2 virus is known to infect the oral cavity and can be readily detected using PCR-based testing. In this study, we examined the host transcriptomic response to PCR-confirmed SARS-CoV-2 virus infection, vaccination against SARS-CoV-2, or breakthrough infection following vaccination using RNA-sequencing. The identification of the viral variant for all samples was obtained using full viral genome sequencing. Approximately equal numbers of males and females were used for every major variant lineage. Results indicate strong anti-viral responses in each case, with some differences due to variant strain and vaccination history, as well as age and sex.
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Project description:Whole blood genome wide DNA methylation profiling from RT-PCR 473 positive and 101 negative SARS-CoV-2 individuals was performed.
2022-07-19 | GSE179325 | GEO
Project description:The COVID-19 pandemic caused by severe acute respiratory syndrome-coronavirus 2 (SARS-CoV-2) has overwhelmed health systems worldwide and highlighted limitations of diagnostic testing. Several types of diagnostic tests including RT-PCR-based assays and antigen detection by lateral flow assays, each with their own strengths and weaknesses, have been developed and deployed in a short time. Here, we describe an immunoaffinity purification approach followed a by high resolution mass spectrometry-based targeted qualitative assay capable of detecting SARS-CoV-2 viral antigen from nasopharyngeal swab samples. Based on our discovery experiments using purified virus, recombinant viral protein and nasopharyngeal swab samples from COVID-19 positive patients, nucleocapsid protein was selected as a target antigen. We then developed an automated antibody capture-based workflow coupled to targeted high-field asymmetric waveform ion mobility spectrometry (FAIMS) - parallel reaction monitoring (PRM) assay on an Orbitrap Exploris 480 mass spectrometer. An ensemble machine learning-based model for determining COVID-19 positive samples was developed using fragment ion intensities from the PRM data. The optimized targeted assay, which was used to analyze 88 positive and 88 negative nasopharyngeal swab samples for validation, resulted in 98% (95% CI = 0.922-0.997) (86/88) sensitivity and 100% (95% CI = 0.958-1.000) (88/88) specificity using RT-PCR-based molecular testing as the reference method. Our results demonstrate that direct detection of infectious agents from clinical samples by tandem mass spectrometry-based assays have potential to be deployed as diagnostic assays in clinical laboratories, which has hitherto been limited to analysis of pure microbial culture
2020-08-13 | PXD020644 | Pride