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.

Unleashing immuno-mass spectrometry superpowers to detect SARS-CoV-2

“…In this article of EBioMedicine, Santosh Renuse and colleagues [1] show the relevance of combining immunoaffinity capture with targeted mass spectrometry measurement to detect SARS-CoV-2 nucleocapsid proteins in nasopharyngeal swab samples. The COVID-19 pandemic has confirmed the need to improve the toolbox available to diagnose respiratory infections.…”

“…The method developed by Renuse and colleagues [1], relies on mass spectrometry, which is fast, sensitive, highly discriminative, and can simultaneously monitor a broad panel of targets. For over a decade now, protein masses have been profiled by whole-cell MALDI-TOF in many microbiological laboratories to instantly identify bacterial isolates grown on agar plates.…”

“…However, these proof-of-concept studies presented data from only few clinical samples. In their report, Renuse and colleagues [1] extend the sensitivity of the approach by introducing SARS-CoV-2 immunocapture and filtering the ions using a differential ion mobility device before mass measurement. They successfully stressed the methodology by testing it on samples with low viral load.…”

“…Since tandem mass spectrometry is already used quite extensively in analytical laboratories in clinical settings, the results presented by Renuse et al [1]. will be significant to clinical scientists.…”

“…Validating this type of methodology on medical samples from large cohorts is the primary objective in the field [10]. Based on the SARS-CoV-2 detection demonstration proposed by Renuse et al [1]. and the continuous cycle of improvements to mass spectrometry instrumentation À regularly increasing sensitivity and throughput À there is no doubt that tandem mass spectrometrybased testing could be the method of choice in future pandemics.…”