Diverse functional autoantibodies in patients with COVID-19

Article activity feed

1. Version published to 10.1038/s41586-021-03631-y

   May 19, 2021
2. 

   ScreenIT

   Mar 1, 2021

   SciScore for 10.1101/2020.12.10.20247205: (What is this?)

   Please note, not all rigor criteria are appropriate for all manuscripts.

   Table 1: Rigor

   Institutional Review Board Statement	IRB: Ethics statement: This study was approved by Yale Human Research Protection Program Institutional Review Boards (FWA00002571, protocol ID 2000027690). Consent: Informed consent was obtained from all enrolled patients and healthcare workers. IACUC: All procedures used in this study (sex-matched, age-matched) complied with federal guidelines and the institutional policies of the Yale School of Medicine Animal Care and Use Committee.
   Randomization	not detected.
   Blinding	Cytokines and FACS analyses were performed blinded.
   Power Analysis	not detected.
   Sex as a biological variable	not detected.
   Cell Line Authentication	not detected.

   Table 2: Resources

   Antibodies
   Sentences	Resou…

   SciScore for 10.1101/2020.12.10.20247205: (What is this?)

   Please note, not all rigor criteria are appropriate for all manuscripts.

   Table 1: Rigor

   Institutional Review Board Statement	IRB: Ethics statement: This study was approved by Yale Human Research Protection Program Institutional Review Boards (FWA00002571, protocol ID 2000027690). Consent: Informed consent was obtained from all enrolled patients and healthcare workers. IACUC: All procedures used in this study (sex-matched, age-matched) complied with federal guidelines and the institutional policies of the Yale School of Medicine Animal Care and Use Committee.
   Randomization	not detected.
   Blinding	Cytokines and FACS analyses were performed blinded.
   Power Analysis	not detected.
   Sex as a biological variable	not detected.
   Cell Line Authentication	not detected.

   Table 2: Resources

   Antibodies
   Sentences	Resources
   Antibodies: Anti-human antibodies used in this study, together with vendors and dilutions, are listed as follows: BB515 anti-hHLA-DR (G46-6) (1:400	Anti-human suggested: None anti-hHLA-DR ( G46-6 ) suggested: None
   Anti-mouse antibodies used in this study, together with vendors and dilutions, are listed as follows: FITC anti-mCD11c (N418) (1:400) (BioLegend), PerCP-Cy5.5 or FITC anti-mLy6C (HK1.4) (1:400) (BioLegend), PE or BV605 or BV711 anti-mNK1.1 (PK136) (1:400) (BioLegend), PE-Cy7 anti-mB220 (RA3-6B2) (1:200) (BioLegend), APC anti-mXCR1 (ZET) (1:200) (BioLegend), APC or AlexaFluor 700 or APC-Cy7 anti-mCD4 (RM4-5) (1:400) (BioLegend), APC-Cy7 anti-mLy6G (1A8) (1:400) (BioLegend), BV605 anti-mCD45 (30-F11) (1:400) (BioLegend), BV711 or PerCP-Cy5.5 anti-mCD8a (53-6.7) (1:400) (BioLegend), AlexaFluor 700 or BV785 anti-mCD11b (M1/70) (1:400) (BioLegend), PE anti-mCXCR3 (CXCR3-173) (1:200) (BioLegend), PE-Cy7 anti-mTCRgd	Anti-mouse suggested: (Santa Cruz Biotechnology Cat# sc-70589, RRID:AB_1120501) anti-mCD11c suggested: None anti-mLy6C suggested: None anti-mNK1.1 suggested: None anti-mB220 ( RA3-6B2 ) suggested: None anti-mXCR1 suggested: None anti-mCD4 suggested: None anti-mLy6G suggested: None anti-mCD45 suggested: None anti-mCD8a suggested: None anti-mCD11b suggested: None anti-mCXCR3 suggested: None CXCR3-173 suggested: None PE-Cy7 suggested: None
   For GPCR N-terminal extracellular domains, these yeast were pooled together with transfected yeast that were used to construct the previously described exoproteome library and a limited dilution of clones were sub-sampled, induced, and stained for FLAG using 1:100 anti-FLAG PE antibody (BioLegend).	anti-FLAG suggested: None
   Plates were washed three times with PBS-T (PBS with 0.1% Tween-20) and 50 μL of HRP anti-Human IgG Antibody at 1:5000 dilution (GenScript) or anti-Human IgM-Peroxidase Antibody at 1:5000 dilution (Sigma-Aldrich) diluted in dilution solution were added to each well.	anti-Human IgG suggested: None anti-Human IgM-Peroxidase suggested: None
   Percent max signal was calculated by subtracting background MFI and calculating values as a percentage of GM-CSF induced pSTAT5 MFI in the absence of IgG. Functional Validation of CXCL1 and CXCL7 Autoantibodies: HTLA cells, a HEK293-derived cell line that stably expresses β-arrestin-TEV and tTA-Luciferase, were seeded in wells of a sterile tissue culture grade flat bottom 96-well plate (35,000 cells/well) in 100 μL DMEM (+ 10% FBS, 1% Penicillin/Streptomycin).	CXCL1 suggested: None CXCL7 suggested: None tTA-Luciferase suggested: None
   Following a wash, cells were blocked with anti-mouse CD16/32 antibodies (BioXCell) for 30 min at 4 °C.	anti-mouse CD16/32 suggested: None
   Experimental Models: Cell Lines
   Sentences	Resources
   Expi293 cells (Thermo Fisher Scientific) were transfected and maintained according to manufacturer protocols.	Expi293 suggested: RRID:CVCL_D615)
   For protein production, Hi-5 cells were infected with P2 virus at a previously optimized titer and harvested 3–5 days after infection.	Hi-5 suggested: None
   Functional Validation of GM-CSF Autoantibodies: TF-1 cells were starved of recombinant GM-CSF (PreproTech, 300-03) eighteen hours prior to experiments.	TF-1 suggested: None
   Experimental Models: Organisms/Strains
   Sentences	Resources
   Mice: B6.Cg-Tg(K18-ACE2)2Prlmn/J (K18-hACE2) mice were purchased from the Jackson Laboratories and were subsequently bred and housed at Yale University.	B6.Cg-Tg(K18-ACE2)2Prlmn/J ( K18-hACE2 ) suggested: None
   Software and Algorithms
   Sentences	Resources
   Ethics statement: This study was approved by Yale Human Research Protection Program Institutional Review Boards (FWA00002571, protocol ID 2000027690).	Yale Human Research Protection Program suggested: None
   The clinical data were collected using EPIC EHR and REDCap 9.3.6 software.	REDCap suggested: (REDCap, RRID:SCR_003445)
   The band corresponding to 257 base pairs was cut out and DNA (NGS library) was extracted using a QIAquick Gel Extraction Kit (Qiagen) according to standard manufacturer protocols.	NGS suggested: (PM4NGS, RRID:SCR_019164)
   SARS-CoV-2 specific antibody ELISA measurement: SARS-CoV-2 specific antibodies were measured as previously described40.	SARS-CoV-2 suggested: (Active Motif Cat# 91351, RRID:AB_2847848)
   Data were analysed using FlowJo software version 10.6 software (Tree Star).	FlowJo suggested: (FlowJo, RRID:SCR_008520)
   Data analysis was performed using R, MATLAB, and GraphPad Prism.	MATLAB suggested: (MATLAB, RRID:SCR_001622) GraphPad Prism suggested: (GraphPad Prism, RRID:SCR_002798)

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   Results from OddPub: We did not detect open data. We also did not detect open code. Researchers are encouraged to share open data when possible (see Nature blog).

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   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.

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   Results from TrialIdentifier: No clinical trial numbers were referenced.

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   Results from Barzooka: We did not find any issues relating to the usage of bar graphs.

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   Results from JetFighter: We did not find any issues relating to colormaps.

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   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.
   • Thank you for including a protocol registration statement.

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   Read the original source
3. 

   Version published to 10.1101/2020.12.10.20247205v5 on medRxiv

   Feb 1, 2021
4. 

   Version published to 10.1101/2020.12.10.20247205v4 on medRxiv

   Dec 19, 2020
5. 

   ScreenIT

   Dec 12, 2020

   SciScore for 10.1101/2020.12.10.20247205: (What is this?)

   Please note, not all rigor criteria are appropriate for all manuscripts.

   Table 1: Rigor

   Institutional Review Board Statement

   Materials and Methods Ethics statement This study was approved by Yale Human Research Protection Program Institutional Review Boards (FWA00002571, protocol ID 2000027690).

   Randomization

   not detected.

   Blinding

   Cytokines and FACS analyses were performed blinded.

   Power Analysis

   not detected.

   Sex as a biological variable

   not detected.

   Cell Line Authentication

   not detected.

   Table 2: Resources

   Antibodies
   Sentences	Resources
   Antibodies Anti-human antibodies used in this study, together with vendors and dilutions, are listed as follows: BB515 anti-hHLA-DR (G46-6) (1:400	Anti-human suggested: None …

   SciScore for 10.1101/2020.12.10.20247205: (What is this?)

   Please note, not all rigor criteria are appropriate for all manuscripts.

   Table 1: Rigor

   Institutional Review Board Statement

   Materials and Methods Ethics statement This study was approved by Yale Human Research Protection Program Institutional Review Boards (FWA00002571, protocol ID 2000027690).

   Randomization

   not detected.

   Blinding

   Cytokines and FACS analyses were performed blinded.

   Power Analysis

   not detected.

   Sex as a biological variable

   not detected.

   Cell Line Authentication

   not detected.

   Table 2: Resources

   Antibodies
   Sentences	Resources
   Antibodies Anti-human antibodies used in this study, together with vendors and dilutions, are listed as follows: BB515 anti-hHLA-DR (G46-6) (1:400	Anti-human suggested: None anti-hHLA-DR ( G46-6 ) suggested: None
   Antimouse antibodies used in this study, together with vendors and dilutions, are listed as follows: FITC anti-mCD11c (N418) (1:400) (BioLegend), PerCP-Cy5.5 or FITC antimLy6C (HK1.4) (1:400) (BioLegend), PE or BV605 or BV711 anti-mNK1.1 (PK136) (1:400) (BioLegend), PE-Cy7 anti-mB220 (RA3-6B2) (1:200) (BioLegend), APC anti-mXCR1 (ZET) (1:200) (BioLegend), APC or AlexaFluor 700 or APC-Cy7 anti-mCD4 (RM4-5) (1:400) (BioLegend), APC-Cy7 anti-mLy6G (1A8) (1:400) (BioLegend), BV605 antimCD45 (30-F11) (1:400) (BioLegend), BV711 or PerCP-Cy5.5 anti-mCD8a (53-6.7) (1:400) (BioLegend), AlexaFluor 700 or BV785 anti-mCD11b (M1/70) (1:400) (BioLegend), PE anti-mCXCR3 (CXCR3-173) (1:200) (BioLegend), PE-Cy7 antimTCRgd	Antimouse suggested: (C. Birchmeier, Max Delbruck Center for Molecular Medicine; Berlin; Germany Cat# Guinea pig anti-mouse Tlx3 polyclonal antibody, RRID:AB_2532145) anti-mCD11c suggested: None antimLy6C suggested: None anti-mNK1.1 suggested: None anti-mB220 ( RA3-6B2 ) suggested: None anti-mXCR1 suggested: None anti-mCD4 suggested: None anti-mLy6G suggested: None antimCD45 suggested: None anti-mCD8a suggested: None anti-mCD11b suggested: None anti-mCXCR3 suggested: None CXCR3-173 suggested: None PE-Cy7 suggested: None
   For GPCR N-terminal extracellular domains, these yeast were pooled together with transfected yeast that were used to construct the previously described exoproteome library and a limited dilution of clones were sub-sampled, induced, and stained for FLAG using 1:100 anti-FLAG PE antibody (BioLegend).	anti-FLAG suggested: None
   Plates were washed three times with PBS-T (PBS with 0.1% Tween-20) and 50 μL of HRP anti-Human IgG Antibody at 1:5000 dilution (GenScript) or anti-Human IgM-Peroxidase Antibody at 1:5000 dilution (Sigma-Aldrich) diluted in dilution solution were added to each well.	anti-Human IgG suggested: None anti-Human IgM-Peroxidase suggested: None
   Percent max signal was calculated by subtracting background MFI and calculating values as a percentage of GM-CSF induced pSTAT5 MFI in the absence of IgG. Functional Validation of CXCL1 and CXCL7 Autoantibodies HTLA cells, a HEK293-derived cell line that stably expresses β-arrestin-TEV and tTALuciferase, were seeded in wells of a sterile tissue culture grade flat bottom 96-well plate (35,000 cells/well) in 100 μL DMEM (+ 10% FBS, 1% Penicillin/Streptomycin).	CXCL1 suggested: None CXCL7 suggested: None
   Following a wash, cells were blocked with anti-mouse CD16/32 antibodies (BioXCell) for 30 min at 4 °C.	anti-mouse CD16/32 suggested: None
   Experimental Models: Cell Lines
   Sentences	Resources
   Expi293 cells (Thermo Fisher Scientific) were transfected and maintained according to manufacturer protocols.	Expi293 suggested: RRID:CVCL_D615)
   For protein production, Hi-5 cells were infected with P2 virus at a previously optimized titer and harvested 3–5 days after infection.	Hi-5 suggested: None
   Experimental Models: Organisms/Strains
   Sentences	Resources
   Mice B6.Cg-Tg(K18-ACE2)2Prlmn/J (K18-hACE2) mice were purchased from the Jackson Laboratories and were subsequently bred and housed at Yale University.	B6.Cg-Tg(K18-ACE2)2Prlmn/J ( K18-hACE2 ) suggested: None
   Software and Algorithms
   Sentences	Resources
   Materials and Methods Ethics statement This study was approved by Yale Human Research Protection Program Institutional Review Boards (FWA00002571, protocol ID 2000027690).	Yale Human Research Protection Program suggested: None
   The clinical data were collected using EPIC EHR and REDCap 9.3.6 software.	REDCap suggested: (REDCap, RRID:SCR_003445)
   The band corresponding to 257 base pairs was cut out and DNA (NGS library) was extracted using a QIAquick Gel Extraction Kit (Qiagen) according to standard manufacturer protocols.	NGS suggested: (PM4NGS, RRID:SCR_019164)
   Data were analysed using FlowJo software version 10.6 software (Tree Star).	FlowJo suggested: (FlowJo, RRID:SCR_008520)
   Data analysis was performed using R, MATLAB, and GraphPad Prism. Reporting Summary Further information on research design will be made available in the Nature Research Reporting Summary linked to this article.	MATLAB suggested: (MATLAB, RRID:SCR_001622) GraphPad suggested: (GraphPad Prism, RRID:SCR_002798)

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   Results from OddPub: Thank you for sharing your data.

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   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.

   ---

   About SciScore

   SciScore is an automated tool that is designed to assist expert reviewers by finding and presenting formulaic information scattered throughout a paper in a standard, easy to digest format. SciScore checks for the presence and correctness of RRIDs (research resource identifiers), and for rigor criteria such as sex and investigator blinding. For details on the theoretical underpinning of rigor criteria and the tools shown here, including references cited, please follow this link.

   Read the original source
6. 

   Version published to 10.1101/2020.12.10.20247205v2 on medRxiv

   Dec 12, 2020
7. 

   Version published to 10.1101/2020.12.10.20247205v3 on medRxiv

   Dec 12, 2020
8. 

   Version published to 10.1101/2020.12.10.20247205v1 on medRxiv

   Dec 11, 2020