Anne-Claude Gingras | Search bioRxiv

bioRxiv2020

A simple protein-based SARS-CoV-2 surrogate neutralization assay

Most of the patients infected with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) mount a humoral immune response to the virus within a few weeks of infection, but the duration of this response and how it correlates with clinical outcomes has not been completely characterized. Of particular importance is the identification of immune correlates of infection that would support public health decision-making on treatment approaches, vaccination strategies, and convalescent plasma therapy. While ELISA-based assays to detect and quantitate antibodies to SARS-CoV-2 in patient samples have been developed, the detection of neutralizing antibodies typically requires more demanding cell-based viral assays. Here, we present a safe and efficient protein-based assay for the detection of serum and plasma antibodies that block the interaction of the SARS-CoV-2 spike protein receptor binding domain (RBD) with its receptor, angiotensin converting-enzyme 2 (ACE2). The assay serves as a surrogate neutralization assay and is performed on the same platform and in parallel with an enzyme-linked immunosorbent assay (ELISA) for the detection of antibodies against the RBD, enabling a direct comparison. The results obtained with our assay correlate with those of two viral based assays, a plaque reduction neutralization test (PRNT) that uses live SARS-CoV-2 virus, and a spike pseudotyped viral-vector-based assay.

immunology↗

bioRxiv2016

Multi-laboratory assessment of reproducibility, qualitative and quantitative performance of SWATH-mass spectrometry

Quantitative proteomics employing mass spectrometry has become an indispensable tool in basic and applied life science research. Methods based on data-dependent acquisition have proved extremely valuable for qualitative proteome analysis but historically have struggled to achieve reproducible quantitative data if large sample cohorts are comparatively analyzed. Targeted proteomics, most commonly implemented as selected reaction monitoring, has emerged as a powerful alternative and succeeded in providing a data independent approach for reproducible quantitative proteomics data but is limited in the number of proteins quantified. SWATH-MS is a recently introduced technique consisting of a data-independent acquisition and a targeted data analysis strategy that aims to maintain the favorable quantitative characteristics (accuracy, sensitivity, specificity) achieved in targeted proteomics but on the scale of thousands of proteins. While previous SWATH-MS studies have shown high intra-lab reproducibility, this has not been evaluated on an inter-lab basis. In this multi-laboratory evaluation study using data from 11 sites worldwide, we have demonstrated that using SWATH-MS we can consistently detect and quantify more than 4,000 proteins from HEK293 cells and that the quantitative protein data generated across laboratories is reproducible. Using synthetic peptide dilution series, we have shown that the sensitivity, dynamic range and reproducibility established with SWATH-MS methods are also uniformly achieved across labs. This study demonstrates that SWATH-MS is a reproducible and accurate technique that can be confidently deployed for large-scale protein quantification in life science research.

Biochemistry↗