Multiplex targeted mass spectrometry assay for one-shot flavivirus diagnosis

Abstract

Targeted proteomic mass spectrometry is emerging as a salient clinical diagnostic tool to track protein biomarkers. However, its strong analytical properties have not been exploited in the diagnosis and typing of flaviviruses. Here, we report the development of a sensitive and specific single-shot robust assay for flavivirus typing and diagnosis using targeted mass spectrometry technology. Our flavivirus parallel reaction monitoring assay (fvPRM) has the ability to track secreted flaviviral nonstructural protein 1 (NS1) over a broad diagnostic and typing window with high sensitivity, specificity, extendibility, and multiplexing capability. These features, pivotal and pertinent to efficient response toward flavivirus outbreaks, including newly emerging flavivirus strains, circumvent the limitations of current diagnostic assays.fvPRM thus carries high potential in positioning itself as a forerunner in delivering early and accurate diagnosis for disease management.

Current and Perspective Diagnostic Techniques for COVID-19

“… 99 There are various techniques for the detection of antigens and antibodies, where a single-shot mass spectrometry can distinguish viral subtypes. 100 A sandwich-type electrochemical immunosensor, which is used for the detection of virus-specific antigen based on a Pd nanoparticle (NP) functionalized electroactive Co-containing metal-organic framework (Co-MOF) for signal amplification, detects viral antigen up to 100 fg/mL. 101 An impedance-based biosensor identifies SARS-CoV-2 pathogens with an ACE2-decorated electrode via a self-assembled monolayer, while the performance usually improves after the insertion of redox-active alkanethiol.…”

Perspective on Proteomics for Virus Detection in Clinical Samples

“…In a publication from 2019, Wee and colleagues published the first parallel reaction monitoring (PRM) assay on an Orbitrap Q Exactive mass spectrometer to detect viruses from serum samples. 25 They were able to identify and differentiate serotypes of DENV from as little as 4 μL of serum. Differentiation of the four serotypes is important in DENV diagnostics because secondary infection with another serotype is associated with severe courses of disease.…”

“…The DENV NS1 protein amounts in serum, for example, range between about 2.5 and 20 pmol/mL. 25 Another example is the HIV p24 antigen amount, which is generally analyzed by ELISA. To obtain CE certification, the LOD for HIV p24 antigen ELISA is 2 IU/mL WHO standard, corresponding to ∼10.3 pg/mL (430 amol/mL).…”

Flavivirus Cross-Reactivity to Dengue Nonstructural Protein 1 Antigen Detection Assays

“…sNS1 is detectable in patient sera as early as the first day of fever and can last even up to nine days after fever onset [7,8]. Therefore, sNS1 can be used as a versatile marker for the early diagnosis of flavivirus infections [9][10][11][12]. However, commercial assays are available only for the detection of DENV NS1 Ag at present.…”