@article{2429afb119374afba7c4ce1e84edcb7a,
title = "A mass spectrometry-based targeted assay for detection of SARS-CoV-2 antigen from clinical specimens",
abstract = "Background: 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. Methods: 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. Findings: 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. Interpretation: 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 cultures.",
keywords = "COVID-19, Diagnostic assays, Ion mobility, Machine learning, Mass spectrometry, SARS-CoV-2",
author = "Santosh Renuse and Vanderboom, {Patrick M.} and Maus, {Anthony D.} and Kemp, {Jennifer V.} and Gurtner, {Kari M.} and Madugundu, {Anil K.} and Sandip Chavan and Peterson, {Jane A.} and Madden, {Benjamin J.} and Mangalaparthi, {Kiran K.} and Mun, {Dong Gi} and Smrita Singh and Kipp, {Benjamin R.} and Surendra Dasari and Singh, {Ravinder J.} and Grebe, {Stefan K.} and Akhilesh Pandey",
note = "Funding Information: The following reagent was deposited by the Centers for Disease Control and Prevention and obtained through BEI Resources, NIAID, NIH: SARS-Related Coronavirus 2, Isolate USA-WA1/2020, Gamma-Irradiated, NR-52287. This study was supported by DBT/Wellcome Trust India Alliance Margdarshi Fellowship grant IA/M/15/1/502023 awarded to AP and the generosity of Eric and Wendy Schmidt. Funding Information: The mass spectrometry raw data have been deposited to the ProteomeXchange Consortium via the PRIDE partner repository with the dataset identifier PXD020644. Skyline files have been deposited to Panorama Public and can be accessed using unique identifier URL (https://panoramaweb.org/covid19ml.url). A.P. S.K.G. R.J.S. B.R.K. S.R. P.M.V. A.D.M designed the study; A.P. S.R. P.M.V. managed the study; S.R. P.M.V. A.D.M, A.P. developed the pipeline of experiment and analysis; A.P. S.R. P.M.V. S.S. D.G.M. performed literature search; S.R. P.M.V, T.D.M. J.V.K. K.M.V. D.G.M. S.C. B.J.M. K.K.M. processed and prepared the samples for LC-MS/MS; J.A.P. performed peptide synthesis; S.R. P.M.V. T.D.M. performed LC-MS/MS; T.D.M. P.M.V. S.D. S.R. A.K.M. processed MS data, performed bioinformatic analysis; S.R. P.M.V. T.D.M. A.K.M. D.G.M. prepared figures and tables with help of other authors; S.R. P.M.V. T.D.M. A.K.M. S.D. A.P. verified the underlaying data; S.R. P.M.V. T.D.M. A.P. wrote the manuscript with help of other authors. All authors read and approved the manuscript. The following reagent was deposited by the Centers for Disease Control and Prevention and obtained through BEI Resources, NIAID, NIH: SARS-Related Coronavirus 2, Isolate USA-WA1/2020, Gamma-Irradiated, NR-52287. This study was supported by DBT/Wellcome Trust India Alliance Margdarshi Fellowship grant IA/M/15/1/502023 awarded to AP and the generosity of Eric and Wendy Schmidt. Publisher Copyright: {\textcopyright} 2021",
year = "2021",
month = jul,
doi = "10.1016/j.ebiom.2021.103465",
language = "English (US)",
volume = "69",
journal = "EBioMedicine",
issn = "2352-3964",
publisher = "Elsevier BV",
}