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Assays for Studying the Role of Vitronectin in Bacterial Adhesion and Serum Resistance
Umeå University, Faculty of Medicine, Department of Molecular Biology (Faculty of Medicine). Clinical Microbiology, Department of Translational Medicine, Lund University.
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2018 (English)In: Journal of Visualized Experiments, ISSN 1940-087X, E-ISSN 1940-087X, no 140, article id e54653Article in journal (Refereed) Published
Abstract [en]

Bacteria utilize complement regulators as a means of evading the host immune response. Here, we describe protocols for evaluating the role vitronectin acquisition at the bacterial cell surface plays in resistance to the host immune system. Flow cytometry experiments identified human plasma vitronectin as a ligand for the bacterial receptor outer membrane protein H of Haemophilus influenzae type f. An enzyme-linked immunosorbent assay was employed to characterize the protein-protein interactions between purified recombinant protein H and vitronectin, and binding affinity was assessed using bio-layer interferometry. The biological importance of the binding of vitronectin to protein H at the bacterial cell surface in evasion of the host immune response was confirmed using a serum resistance assay with normal and vitronectin-depleted human serum. The importance of vitronectin in bacterial adherence was analyzed using glass slides with and without vitronectin coating, followed by Gram staining. Finally, bacterial adhesion to human alveolar epithelial cell monolayers was investigated. The protocols described here can be easily adapted to the study of any bacterial species of interest.

Place, publisher, year, edition, pages
Cambride, USA: Journal of Visualized Experiments , 2018. no 140, article id e54653
Keywords [en]
Immunology and Infection, Issue 140, Adhesion, Bacteria, Epithelial Cells, Infection, Protein-Protein teraction, Respiratory Tract Pathogen, Serum Resistance Assay, Vitronectin
National Category
Immunology in the medical area Microbiology in the medical area
Identifiers
URN: urn:nbn:se:umu:diva-156339DOI: 10.3791/54653ISI: 000456452800109PubMedID: 30394376OAI: oai:DiVA.org:umu-156339DiVA, id: diva2:1288129
Available from: 2019-02-12 Created: 2019-02-12 Last updated: 2019-02-12Bibliographically approved

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Singh, Birendra

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CiteExportLink to record
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