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They can improve bacterial survival, mucosal retention and therapeutic delivery, but the structural basis of these effects is often unclear. This review groups biointerfaces by their dominant role: barrier protection, recognition, responsiveness, interaction and renewal by living cells. We compare microscopy methods for measuring coating continuity, ligand organisation, surface mechanics and changes during growth or biological transport. We also explain how artificial intelligence (AI)\u2010assisted image analysis can convert bacterial images into reproducible single\u2010cell measurements, using published workflows as practical examples. These methods can help connect interfacial changes with mucosal protection, immune regulation, targeted delivery and antibiofilm activity. Coating continuity, ligand organisation and mechanics are parameters associated with performance; their causal contributions remain to be established. We therefore propose combining measurements over time with matched functional assays and controlled changes to individual interface parameters. This approach can test structure\u2013property\u2013function relationships and guide the design of reproducible living therapeutics.<\/jats:p>","DOI":"10.1111\/jmi.70182","type":"journal-article","created":{"date-parts":[[2026,9,29]],"date-time":"2026-09-29T10:57:04Z","timestamp":1790679424000},"update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Engineered probiotic biointerfaces: Construction principles, microscopic characterisation and biomedical functions"],"prefix":"10.1111","author":[{"given":"Jiaming","family":"Tang","sequence":"first","affiliation":[{"name":"School of Materials Science and Engineering Institute for Advanced Materials Jiangsu University  Zhenjiang China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Mengqi","family":"Bai","sequence":"additional","affiliation":[{"name":"Department of Laboratory Medicine, School of Medicine Jiangsu University  Zhenjiang Jiangsu 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