PMC:7108609 / 11162-14673
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{"target":"https://pubannotation.org/docs/sourcedb/PMC/sourceid/7108609","sourcedb":"PMC","sourceid":"7108609","source_url":"https://www.ncbi.nlm.nih.gov/pmc/7108609","text":"Inhibition of adhesion by anti-A antibodies\nThe effect of anti-A antibodies on the S protein/ACE2 interaction was first tested using a monoclonal anti-A. A clear-cut inhibition of the cell adhesion was observed using the monoclonal antibody 3-3A at 2 μg/mL. Specificity of the inhibition was confirmed since a control irrelevant antibody failed to inhibit and the adhesion to Vero cells of S protein-transfected CHO cells lacking the A antigen was not inhibited by the anti-A mAb (Figure 2D). Vero cells do not express the A histo-blood group antigen. Therefore, the inhibition of adhesion mediated by the anti-A mAb can only result from a binding to CHO S protein expressing cells and not to the glycans of ACE2. In order to assay the ability of natural human anti-A to inhibit cell adhesion, plasma samples from two O blood group individuals with high anti-A titers were selected. The samples were first adsorbed on silica beads conjugated to the A type 2 tetrasaccharide in order to specifically remove the anti-A natural plasma antibodies. Efficacy of the adsorption was controlled by ELISA which showed that the reactivity to the A type 2 tetrasaccharide was almost completely abolished following adsorption (Figure 3B). The A type 2 adsorbed and mock adsorbed plasma samples from the two individuals were then added in the cell-based assay. Both mock adsorbed samples, containing the anti-A as shown in Figure 3B, strongly inhibited the adhesion of A antigen-S protein expressing cells to Vero cells. In both cases, this inhibition was almost completely lost after A type 2 adsorption, showing that it was specifically mediated by anti-A plasma antibodies (Figure 3C). Moreover, the inhibition of adhesion by blood group O plasma was dose-dependent and still detected at a plasma dilution as low as 1/32 (Figure 3D).\nFig. 3 Effect of anti-A antibodies on the interaction between the SARS-CoV S protein and ACE2. (A) The anti-A monoclonal antibody 3-3A was added at the indicated concentrations to the triple transfected CHO cells suspension prior to incubation on the Vero cell layer. An irrelevant IgG1 was used as control at 4 μg/mL. The results are presented as mean cell number per field ±SD of one representative experiment out of two. From 1.0 μg/mL to 4.0 μg/mL anti-A, values are significantly different from those for the control IgG (P \u003c 0.05 and 0.001, respectively). (B) Adsorption of the anti-A natural antibody from human O plasmas. Plasma samples from two individuals were adsorbed on either control silica beads (mock) or A type 2 tetrasaccharide conjugated to silica beads (At2). The postadsorption plasma reactivity on A type 2 conjugated to polyacrylamide was tested by ELISA. Results are shown as O.D. 450 nm values of duplicate wells ±SD for each plasma sample diluted at 1/4. In the absence of A type 2 conjugate, mean O.D. values were 0.13. (C) Inhibition of the adhesion of CHO triple transfected cells to Vero cells by mock adsorbed (mock) or A type 2 (At2) adsorbed human blood group O plasma samples from individuals 1 and 2. Plasma samples were diluted at 1/8 in PBS. Control values were obtained in the absence of plasma. Values for the mock adsorbed plasma were significantly different from the control values (P \u003c 0.001). (D) Inhibition of the adhesion in the cell-based assay as in C by serial dilutions of unadsorbed plasma from individual 1. All values obtained in the presence of plasma were significantly different from the control value (from P \u003c 0.05 to P \u003c 0.0001).","divisions":[{"label":"title","span":{"begin":0,"end":43}},{"label":"p","span":{"begin":44,"end":1822}},{"label":"label","span":{"begin":1823,"end":1829}}],"tracks":[]}