Public Health: Add mass spectrometry to the pandemic toolbox
A new protocol step improves robustness and ease-of-use for mass spectrometry in the clinic, opening the door to mass deployment to monitor infectious agents.
- Laboratory of Pharmaceutical Biotechnology, Ghent University, Belgium
- ProGenTomics, Belgium
Widespread testing has become a cornerstone of the response against the COVID-19 pandemic. So far, this has almost exclusively been done by detecting the genetic information of the SARS-CoV-2 virus (its mRNA) using tests such as RT-PCR. This approach is efficient, relatively simple and acceptably cheap. However, only relying on one type of technology can lead to supply shortages, and it makes it difficult to assess how well the method fares in terms of sensitivity, false positives and false negatives (Woolston, 2021). Additionally, while RT-PCR tests can assess whether someone is carrying the virus, they cannot reveal how many viral particles someone is releasing into the environment, how infectious a person is, or how a patient will fare. The pandemic ‘readiness toolbox’ must therefore be extended to include methods that can detect other types of biomolecules beyond mRNA, such as viral proteins and peptides (Evans et al., 2021).
Mass spectrometry, an approach that helps to assess which compounds are present in a sample, is an obvious choice. Rather than detecting a precise target (like RT-PCR assays do with mRNA), this intrinsically versatile method measures the physical properties of any and many biomolecules, including peptides derived from proteins. Mass spectrometry can therefore monitor a practically limitless number of molecules, making it a sustainable analytical technique. In fact, an instrument calibrated to quantify certain disease-related proteins in plasma, for example, can start measuring peptides derived from a new pathogen in next to no time (Grossegesse et al., 2020; Van Puyvelde et al., 2021).
However, despite the high-level sensitivity and accuracy of mass spectrometry, clinical settings often rely on other techniques to analyze proteins, as extensive expertise is believed to be needed both to handle the instrument and to interpret the data. Most of these concerns are due the additional, meaningless data from all the other proteins and molecules in the sample (‘the noise’), which complicate the detection of the peptides of interest (the actual signal). Indeed, like any other analytical technique, mass spectrometry measures the signal-to-noise ratio, but the instruments cannot automatically reduce the noise from this equation. This has made mass spectrometry a frightening prospect for clinical implementation, reducing instrumental robustness and complicating data interpretation. In response, scientists usually strive to improve the signal-to-noise ratio by increasing the signal. Now, in eLife, Fredrik Edfors and colleagues at institutions in Sweden and Canada – including Andreas Hober as first author – report having tweaked how to prepare a mass spectrometry sample to prevent background noise from emerging instead (Hober et al., 2021).
The team added a new step in the protocol, called peptide immuno-enrichment, which involves attaching antibodies to magnetic beads to ‘fish’ the target peptides directly out of the patient samples before analysis (Razavi et al., 2016; Anderson et al., 2004). This multiplies the sensitivity many-fold and reduces measurement time, interferences and instrument contamination while also increasing robustness (as also proposed in Van Puyvelde et al., 2021). Additionally, without the noise, mass spectrometry can determine the amount of a given target protein extremely accurately, something that is not possible using RT-PCR (Evans et al., 2021).
In other words, Hober et al. have set the stage for applying mass spectrometry to accurately quantify SARS-CoV-2 peptides in a variety of patient samples. With this approach, each instrument could process samples from 500 patients in a single day. In fact, with peptide immuno-enrichment already used to assess a panel of inflammation proteins in plasma in the clinic, adding an antibody bead targeting a peptide derived from a pathogen does not change the overall workflow (Anderson et al., 2020). The complete sample preparation protocol with peptide immuno-enrichment could be done for less than 40€ per sample, in less than three hours of automatable sample preparation and without requiring any mass spectroscopy expertise to interpret the data. Beyond this versatility, another important characteristic of the approach proposed by Hober et al. is that the ‘fishing’ of the target peptides theoretically means that patient samples could be pooled and analysed together. As suggested for RT-PCR (Verwilt et al., 2020), this strategy would increase throughput and save resources.
Taken together, these features provide an opportunity to develop an early warning test that targets a dozen respiratory pathogens in pools of up to 32 patient samples, with each machine being able to run 500 pooled samples – totaling up to 16,000 individuals a day. Doing this throughout the year would help to monitor how a pathogen spreads through a population over time. In fact, if several pathogens start to pass through a population simultaneously, other mass spectrometry instruments can easily be set up to detect these infectious agents in parallel in unpooled samples (up to 500 patients a day per instrument).
The question then becomes: should we now start using mass spectrometry instead of RT-PCR in the present COVID-19 pandemic? Currently, precisely assessing the amount of viral proteins may not help to decide whether a person should be quarantined. However, this information may help to decide when an individual should be released into the community as viral protein might more closely track infectivity in SARS-CoV-2 than viral mRNA, which remains detectable after a patient ceases to be infectious (Evans et al., 2021). Irrespectively, starting to measure viral protein loads in thousands of COVID-19 patients using mass spectrometry could help the technique to mature into a reliable approach to add to the pandemic readiness toolbox; in this effort, the work by Hober et al. will help revolutionize the way mass spectrometry is approached in the clinic.
References
-
Perspective on proteomics for virus detection in clinical samplesJournal of Proteome Research 19:4380–4388.https://doi.org/10.1021/acs.jproteome.0c00674
Article and author information
Author details
Publication history
Copyright
© 2021, Van Puyvelde and Dhaenens
This article is distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use and redistribution provided that the original author and source are credited.
Metrics
-
- 935
- views
-
- 50
- downloads
-
- 6
- citations
Views, downloads and citations are aggregated across all versions of this paper published by eLife.
Download links
A two-part list of links to download the article, or parts of the article, in various formats.
Downloads (link to download the article as PDF)
Open citations (links to open the citations from this article in various online reference manager services)
Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)
Public Health: Add mass spectrometry to the pandemic toolbox
eLife 10:e75471.
https://doi.org/10.7554/eLife.75471
Further reading
-
- Immunology and Inflammation
The incidence of metabolic dysfunction-associated steatotic liver disease (MASLD) has been increasing worldwide. Since gut-derived bacterial lipopolysaccharides (LPS) can travel via the portal vein to the liver and play an important role in producing hepatic pathology, it seemed possible that (1) LPS stimulates hepatic cells to accumulate lipid, and (2) inactivating LPS can be preventive. Acyloxyacyl hydrolase (AOAH), the eukaryotic lipase that inactivates LPS and oxidized phospholipids, is produced in the intestine, liver, and other organs. We fed mice either normal chow or a high-fat diet for 28 weeks and found that Aoah-/- mice accumulated more hepatic lipid than did Aoah+/+ mice. In young mice, before increased hepatic fat accumulation was observed, Aoah-/- mouse livers increased their abundance of sterol regulatory element-binding protein 1, and the expression of its target genes that promote fatty acid synthesis. Aoah-/- mice also increased hepatic expression of Cd36 and Fabp3, which mediate fatty acid uptake, and decreased expression of fatty acid-oxidation-related genes Acot2 and Ppara. Our results provide evidence that increasing AOAH abundance in the gut, bloodstream, and/or liver may be an effective strategy for preventing or treating MASLD.
-
- Immunology and Inflammation
- Microbiology and Infectious Disease
Pseudomonas aeruginosa (PA) is an opportunistic, frequently multidrug-resistant pathogen that can cause severe infections in hospitalized patients. Antibodies against the PA virulence factor, PcrV, protect from death and disease in a variety of animal models. However, clinical trials of PcrV-binding antibody-based products have thus far failed to demonstrate benefit. Prior candidates were derivations of antibodies identified using protein-immunized animal systems and required extensive engineering to optimize binding and/or reduce immunogenicity. Of note, PA infections are common in people with cystic fibrosis (pwCF), who are generally believed to mount normal adaptive immune responses. Here, we utilized a tetramer reagent to detect and isolate PcrV-specific B cells in pwCF and, via single-cell sorting and paired-chain sequencing, identified the B cell receptor (BCR) variable region sequences that confer PcrV-specificity. We derived multiple high affinity anti-PcrV monoclonal antibodies (mAbs) from PcrV-specific B cells across three donors, including mAbs that exhibit potent anti-PA activity in a murine pneumonia model. This robust strategy for mAb discovery expands what is known about PA-specific B cells in pwCF and yields novel mAbs with potential for future clinical use.
