No change is seen in the infection-naive group against the Wuhan Hu-1 bait, but an increase is observed for the pre-infection group at first vaccination

No change is seen in the infection-naive group against the Wuhan Hu-1 bait, but an increase is observed for the pre-infection group at first vaccination. may better reflect neutralization to VoCs. Analyzing additional immunoproteins beyond immunoglobulin (Ig) G, provides important information about our understanding of the response to infection and vaccination. Keywords:complement: immunoglobulin, SARS-CoV-2, vaccination, proteomics, COVID-19, variant of concern, mass spectrometry, omicron variant, delta variant == Graphical abstract == == Highlights == Multiplexed targeted proteomics is a viable alternative to ELISA-based testing The immunocomplex to SARS-CoV-2 spike is altered against variants of concern (VoCs) IgA is an indicator of prior SARS-CoV-2 infection C1q can reflect neutralization against VoCs better than IgG1 == Motivation == Assays for measuring serum antibody responses are typically limited to measurement of a total or single immunoglobulin isotype. The antibody response is far more complex, with multiple immunoglobulin classes, isotypes, and complement factors involved. AM679 This is a potential wealth of information that is typically understudied and missed by existing tests. The global COVID-19 pandemic has highlighted the need to understand better the immune response in respect to vaccine AM679 development and emerging new severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) variants. Using the ability of tandem mass spectrometry to multiplex and directly and accurately measure the antibody complex, we devised an alternative assay to capture this valuable information. Doykov et al. present an assay that reveals changes in immunoglobulin classes, subtypes, and complement in response to vaccination, infection, and variants of concern. The assay can provide insight into an individuals immune response and aid vaccine design and delivery. It can also be applied to other antigens and diseases. == Introduction == After the first cases of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) were identified in late 2019, 20202021 saw the development and rollout of the worlds fastest and largest global vaccination programs. However, with potential waning immunity over time (Gaebler et al., 2021) and the impact of infection from emerging variants of concern (VoCs) (Reynolds et al., 2021a), it is apparent that there is a need for better and more informative testing (Abbasi, 2021). This will help determine the clinical need for booster vaccination and timing of the boost itself. First-generation tests were rolled out at scale but are largely based on simple nonspecific binding to the prototypic Wuhan Hu-1 spike sequence. It is now clear that these methods overestimate the AM679 actual protective immunity against VoCs (Reynolds et al., 2021a,2021b,2022). Current technologies that directly measure accepted correlates of protection such as neutralizing antibodies (nAbs) scale poorly for clinical utility, while serological approaches (ELISA or electrochemiluminescent immunoassay [ECLIA]) measure only part of the antibody response and omit measurement of effector Fc antibody functions such as complement involvement (Ju et al., 2020;Nie et al., 2020;Yu et al., 2020). To aid in understanding the antibody response to SARS-CoV-2, we have developed a methodology that includes a AM679 bait and capture system, followed by a multiplexed and targeted proteomic liquid chromatography-tandem mass spectrometry (LC-MS/MS) analyses. The combination of immunocapture with the multiplexing capability to look at multiple proteins involved in the immune response and high accuracy of mass Flt1 spectrometry quantitation makes this an extremely powerful and more informative combination. In addition, tandem mass spectrometers are also routinely used for small molecule clinical assays in most UK pathology laboratories and are therefore platforms that could be utilized for targeted proteomic assays. It is only recently, with improving technology, that they are becoming recognized for their potential clinical application for multiplex protein analysis (Smit et al., 2021). In this work, we describe how we have used this assay to compare with previously determined immune correlates (Reynolds et al., 2021a,2021b,2022) in serial samples, in response to vaccination, infection, and an individuals potential protection against VoCs. This analysis was performed using serum samples from the COVIDsortium study (Manisty et al., 2021a,2021b;Reynolds et.