A streamlined whole blood CyTOF workflow defines a circulating immune cell signature of COVID ‐19
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Abstract
Mass cytometry (CyTOF) represents one of the most powerful tools in immune phenotyping, allowing high throughput quantification of over 40 parameters at single‐cell resolution. However, wide deployment of CyTOF‐based immune phenotyping studies are limited by complex experimental workflows and the need for specialized CyTOF equipment and technical expertise. Furthermore, differences in cell isolation and enrichment protocols, antibody reagent preparation, sample staining, and data acquisition protocols can all introduce technical variation that can confound integrative analyses of large data‐sets of samples processed across multiple labs. Here, we present a streamlined whole blood CyTOF workflow which addresses many of these sources of experimental variation and facilitates wider adoption of CyTOF immune monitoring across sites with limited technical expertise or sample‐processing resources or equipment. Our workflow utilizes commercially available reagents including the Fluidigm MaxPar Direct Immune Profiling Assay (MDIPA), a dry tube 30‐marker immunophenotyping panel, and SmartTube Proteomic Stabilizer, which allows for simple and reliable fixation and cryopreservation of whole blood samples. We validate a workflow that allows for streamlined staining of whole blood samples with minimal processing requirements or expertise at the site of sample collection, followed by shipment to a central CyTOF core facility for batched downstream processing and data acquisition. We apply this workflow to characterize 184 whole blood samples collected longitudinally from a cohort of 72 hospitalized COVID‐19 patients and healthy controls, highlighting dynamic disease‐associated changes in circulating immune cell frequency and phenotype.
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SciScore for 10.1101/2020.06.26.20141341: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Institutional Review Board Statement IRB: Samples: Samples were collected from consented healthy donors and hospitalized COVID-19 patients under protocols approved by the Mount Sinai Institutional Review Board. Randomization not detected. Blinding not detected. Power Analysis not detected. Sex as a biological variable not detected. Table 2: Resources
Software and Algorithms Sentences Resources Data analysis: Debarcoded files were uploaded to Cytobank for analysis. Cytobanksuggested: (Cytobank, RRID:SCR_014043)In some cases, immune cell populations were also identified using automated approaches including MaxPar PathSetter and Astrolabe, the result of which largely correlated well with our manual gating … SciScore for 10.1101/2020.06.26.20141341: (What is this?)
Please note, not all rigor criteria are appropriate for all manuscripts.
Table 1: Rigor
Institutional Review Board Statement IRB: Samples: Samples were collected from consented healthy donors and hospitalized COVID-19 patients under protocols approved by the Mount Sinai Institutional Review Board. Randomization not detected. Blinding not detected. Power Analysis not detected. Sex as a biological variable not detected. Table 2: Resources
Software and Algorithms Sentences Resources Data analysis: Debarcoded files were uploaded to Cytobank for analysis. Cytobanksuggested: (Cytobank, RRID:SCR_014043)In some cases, immune cell populations were also identified using automated approaches including MaxPar PathSetter and Astrolabe, the result of which largely correlated well with our manual gating approaches. MaxPar PathSettersuggested: NoneResults from OddPub: Thank you for sharing your data.
Results from LimitationRecognizer: An explicit section about the limitations of the techniques employed in this study was not found. We encourage authors to address study limitations.Results from TrialIdentifier: No clinical trial numbers were referenced.
Results from Barzooka: We did not find any issues relating to the usage of bar graphs.
Results from JetFighter: We did not find any issues relating to colormaps.
Results from rtransparent:- Thank you for including a conflict of interest statement. Authors are encouraged to include this statement when submitting to a journal.
- Thank you for including a funding statement. Authors are encouraged to include this statement when submitting to a journal.
- No protocol registration statement was detected.
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