Secretory IgA and T cells targeting SARS-CoV-2 spike protein are transferred to the breastmilk upon mRNA vaccination

Article activity feed

1. Version published to 10.1016/j.xcrm.2021.100468

   Dec 1, 2021
2. 

   ScreenIT

   May 7, 2021

   SciScore for 10.1101/2021.05.03.21256416: (What is this?)

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

   Table 1: Rigor

   …

   Ethics	Consent: All participants provided informed consent and all procedures were approved by NOVA Medical School ethics committee, in accordance with the provisions of the Declaration of Helsinki and the Good Clinical Practice guidelines of the International Conference on Harmonization. IRB: All participants provided informed consent and all procedures were approved by NOVA Medical School ethics committee, in accordance with the provisions of the Declaration of Helsinki and the Good Clinical Practice guidelines of the International Conference on Harmonization.
   Sex as a biological variable	not detected.
   Randomization	not detected.
   Blinding	not detected.
   Power Analysis	not detected.
   Cell Line Authentication

   SciScore for 10.1101/2021.05.03.21256416: (What is this?)

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

   Table 1: Rigor

   Ethics	Consent: All participants provided informed consent and all procedures were approved by NOVA Medical School ethics committee, in accordance with the provisions of the Declaration of Helsinki and the Good Clinical Practice guidelines of the International Conference on Harmonization. IRB: All participants provided informed consent and all procedures were approved by NOVA Medical School ethics committee, in accordance with the provisions of the Declaration of Helsinki and the Good Clinical Practice guidelines of the International Conference on Harmonization.
   Sex as a biological variable	not detected.
   Randomization	not detected.
   Blinding	not detected.
   Power Analysis	not detected.
   Cell Line Authentication	not detected.

   Table 2: Resources

   Antibodies
   Sentences	Resources
   PBMCs were stained with a fixable viability dye eFluor™ 506 (invitrogen) and surface labelled with the following antibodies all from BioLegend: anti-CD3 (UCHT1), anti-CD4 (SK3), anti-OX40 (Ber-ACT35)	anti-CD3 suggested: None UCHT1 suggested: None anti-CD4 suggested: None anti-OX40 suggested: None
   ELISA: Antibody binding to SARS-CoV-2 trimeric spike protein or its RBD domain was assessed by a previously described in-house ELISA assay 42 based on the protocol by Stadlbauer et al 66.	SARS-CoV-2 trimeric spike protein suggested: None
   Plates were washed and in incubated for 30 min at room temperature with 1:25,000 dilution of HRP-conjugated anti-human IgA, IgG and IgM antibodies (Abcam, ab97225/ab97215/ab97205	anti-human IgA , IgG suggested: None IgM suggested: (Abcam Cat# ab97205, RRID:AB_10695942)
   goat anti-human IgA/IgG/IgM-HRP secondary antibodies (diluted at in 1% BSA-0.05% PBS-T.	anti-human IgA/IgG/IgM-HRP suggested: None
   Western blots (WB) were performed using nitrocellulose membranes for gel transfer and goat anti-human IgA alpha chain (HRP) (Abcam) with SuperSignal™ West Pico Chemiluminescent Substrate (ThermoFischer Scientific) for antibody detection, in Trans-Blot Turbo Transfer System (Bio-Rad Laboratories, Inc)	anti-human IgA alpha chain ( HRP suggested: None
   Experimental Models: Cell Lines
   Sentences	Resources
   Production of 293T cells stably expressing human ACE2 receptor: Production of 293T cells stably expressing human ACE2 receptor was done as previously described 43.	293T suggested: None
   Briefly, VSV-G pseudotyped lentiviruses encoding human ACE2, 293ET cells were transfected with pVSV-G, psPAX2 and pLEX-ACE2 using jetPRIME (Polyplus), according to manufacturer’s instructions.	293ET suggested: RRID:CVCL_6996)
   The 293T-Ace2 cell line was passaged six times before use and kept in culture medium supplemented with 1.25 μg/ml puromycin.	293T-Ace2 suggested: RRID:CVCL_YZ65)
   Recombinant DNA
   Sentences	Resources
   Briefly, VSV-G pseudotyped lentiviruses encoding human ACE2, 293ET cells were transfected with pVSV-G, psPAX2 and pLEX-ACE2 using jetPRIME (Polyplus), according to manufacturer’s instructions.	pVSV-G suggested: RRID:Addgene_138479) psPAX2 suggested: RRID:Addgene_12260) pLEX-ACE2 suggested: None
   Production and titration of spike pseudotyped lentiviral particles: To generate spike pseudotyped lentiviral particles, 6×106 293ET cells were co-transfected with 8.89ug pLex-GFP reporter, 6.67μg psPAX2, and 4.44μg pCAGGS-SARS-CoV-2-Strunc D614G, using jetPRIME according to manufacturer’s instructions.	pLex-GFP suggested: None pCAGGS-SARS-CoV-2-Strunc D614G suggested: None
   Software and Algorithms
   Sentences	Resources
   Cells were washed, fixed with 1% PFA and acquired in BD FACS Aria III equipment (BD Biosciences) and analysed with FlowJo v10.7.3 software (Tree Star).	FlowJo suggested: (FlowJo, RRID:SCR_008520)
   Western blots (WB) were performed using nitrocellulose membranes for gel transfer and goat anti-human IgA alpha chain (HRP) (Abcam) with SuperSignal™ West Pico Chemiluminescent Substrate (ThermoFischer Scientific) for antibody detection, in Trans-Blot Turbo Transfer System (Bio-Rad Laboratories, Inc)	Bio-Rad Laboratories suggested: (Bio-Rad Laboratories, RRID:SCR_008426)
   Briefly, VSV-G pseudotyped lentiviruses encoding human ACE2, 293ET cells were transfected with pVSV-G, psPAX2 and pLEX-ACE2 using jetPRIME (Polyplus), according to manufacturer’s instructions.	Polyplus suggested: None
   Statistical analysis: The half-maximal neutralization titre (NT50), defined as the reciprocal of the dilution at which infection was decreased by 50%, was determined using four-parameter nonlinear regression (least squares regression without weighting; constraints: bottom=0) (GraphPad Prism 9).	GraphPad suggested: (GraphPad Prism, RRID:SCR_002798)

   ---

   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).

   ---

   Results from LimitationRecognizer: We detected the following sentences addressing limitations in the study:

   These results highlight the functional compartmentalization between mucosal and the systemic immunity and the limitations of systemic administered BNT162b2 vaccine in eliciting a strong mucosal response 54. The function of antibodies produced within the mammary gland MALT is to provide protective immunity to the suckling infant through the secretion of polymeric antibodies complexed to j-chain and secretory component proteins 13. The secretory component is essential to ensure that milk antibodies are effectively transferred via the breastmilk by providing protection from proteolytic cleavage and by facilitating systemic uptake and tissue distribution in the infant 13. Spike-reactive SIgA were detected in the breast milk of COVID-19 patients 55, nonetheless whether SIgA was similarly present in the breast milk upon mRNA vaccination had remained unaddressed. By combining size exclusion chromatography with customized spike-ELISA, we concluded that vaccination does in fact originate spike-reactive SIgA, which is secreted predominately as tetramers. We detected SIgA in 57% of milk samples. Our detected milk SIgA prevalence is on the lower-end of the interval of milk IgA prevalence (61.8% to >80%) detected by others 16-20. This discrepancy can probably be ascribed to different experimental procedures. While we detected SIgA in milk samples that had been diluted 50-fold, these previous works either used undiluted milk samples or diluted them at a maximum of 5-fold. Upon administrati...

   ---

   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.

   ---

   Results from scite Reference Check: We found no unreliable references.

   ---

   Read the original source
3. 

   Version published to 10.1101/2021.05.03.21256416 on medRxiv

   May 4, 2021