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. 2021 Nov 23;11:22799. doi: 10.1038/s41598-021-02065-w

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© The Author(s) 2021

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.

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IsdB-mediated adhesion to HUVEC cell monolayers. (A) Confluent umbilical vein endothelial cell (HUVEC) monolayers were incubated with the calcium ionophore A23187 and fixed with paraformaldehyde. The monolayers were incubated with the indicated amounts of IsdB NEAT1-NEAT2 and binding of the ligand to the cells was detected by addition to the wells of a rabbit polyclonal IsdB antibody followed by an HRP-conjugated goat anti-rabbit IgG. The binding of IsdB NEAT1-NEAT2 to the ionophore-untreated cells is also reported. Statistically significant difference is indicated (***, P < 0.001). (B) Endothelial cell monolayers treated as reported in A were incubated with S. aureus SH1000 WT and isdB mutant bacteria. Adherence to the monolayers was detected by the addition of HRP-conjugated rabbit anti-mouse IgG to the wells. The effect of antibodies on the adhesion of bacteria to endothelial cells was tested by incubating S. aureus cells with monolayers in the presence of an anti-A1 6D1 antibody or an isotype control mAb. Binding of HRP-conjugated rabbit anti-mouse IgG to monolayers in the absence of bacteria is also reported. Statistically significant difference is indicated (***, P < 0.001). Error bars show S.D. of the means from three independent determinations, each performed in triplicate.