{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,21]],"date-time":"2026-10-21T23:17:10Z","timestamp":1792624630176,"version":"4.5.0"},"reference-count":25,"publisher":"Wiley","issue":"6","license":[{"start":{"date-parts":[[2022,6,7]],"date-time":"2022-06-07T00:00:00Z","timestamp":1654560000000},"content-version":"vor","delay-in-days":6,"URL":"http:\/\/creativecommons.org\/licenses\/by-nc-nd\/4.0\/"}],"funder":[{"DOI":"10.13039\/100000060","name":"National Institute of Allergy and Infectious Diseases","doi-asserted-by":"publisher","award":["R21 AI163816"],"award-info":[{"award-number":["R21 AI163816"]}],"id":[{"id":"10.13039\/100000060","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100000060","name":"National Institute of Allergy and Infectious Diseases","doi-asserted-by":"publisher","award":["R21 AI149180"],"award-info":[{"award-number":["R21 AI149180"]}],"id":[{"id":"10.13039\/100000060","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100000060","name":"National Institute of Allergy and Infectious Diseases","doi-asserted-by":"publisher","award":["R01 AI151029"],"award-info":[{"award-number":["R01 AI151029"]}],"id":[{"id":"10.13039\/100000060","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100000060","name":"National Institute of Allergy and Infectious Diseases","doi-asserted-by":"publisher","award":["U01 AI150748"],"award-info":[{"award-number":["U01 AI150748"]}],"id":[{"id":"10.13039\/100000060","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100000050","name":"National Heart, Lung, and Blood Institute","doi-asserted-by":"publisher","award":["R01 HL151840"],"award-info":[{"award-number":["R01 HL151840"]}],"id":[{"id":"10.13039\/100000050","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Current Protocols"],"published-print":{"date-parts":[[2022,6]]},"abstract":"<jats:title>Abstract<\/jats:title><jats:p>A diverse collection of viral pathogens target airway epithelial cells for infection, with effects ranging from mild upper respiratory tract symptoms to death of the infected individual. Among these pathogens are recently discovered and\/or emergent viruses that sometimes fail to infect commonly used, immortalized cell lines and for which infection phenotypes in the respiratory tract remain unknown. Human airway epithelial cultures have been developed over the past several decades and have proven to be a useful model system in culturing hard\u2010to\u2010grow viruses and assaying various features of infection in a physiologically relevant setting. This article includes methods for the generation of well\u2010differentiated human airway epithelial cell cultures at air\u2010liquid interface that recapitulate the mucosal epithelium of the trachea\/bronchus <jats:italic>in vivo<\/jats:italic>. We further detail inoculation of these cultures with respiratory viruses\u2014specifically rhinovirus, influenza virus, and severe acute respiratory syndrome coronavirus 2 (SARS\u2010CoV\u20102)\u2014and provide a protocol for the detection of double\u2010stranded RNA or viral antigen\u2013positive cells by immunofluorescence microscopy. These techniques, together with a post\u2010imaging analysis, can be applied to characterize the efficiency of infection and kinetics of spread within the airway epithelium. Furthermore, these methods can be utilized in conjunction with antibodies against cellular targets to determine cell tropism and colocalization with specific host factors during infection. \u00a9 2022 The Authors. Current Protocols published by Wiley Periodicals LLC.<\/jats:p><jats:p><jats:bold>Basic Protocol 1<\/jats:bold>: Generation of human airway epithelial cultures at air\u2010liquid interface (HAE\u2010ALI)<\/jats:p><jats:p><jats:bold>Basic Protocol 2<\/jats:bold>: Viral inoculation of HAE\u2010ALI<\/jats:p><jats:p><jats:bold>Basic Protocol 3<\/jats:bold>: Immunofluorescence (IF)\u2010based detection of infected cells in HAE\u2010ALI<\/jats:p>","DOI":"10.1002\/cpz1.453","type":"journal-article","created":{"date-parts":[[2022,6,7]],"date-time":"2022-06-07T16:26:31Z","timestamp":1654619191000},"source":"Crossref","is-referenced-by-count":13,"title":["Immunofluorescence\u2010Mediated Detection of Respiratory Virus Infections in Human Airway Epithelial Cultures"],"prefix":"10.1002","volume":"2","author":[{"given":"Talita B.","family":"Gagliardi","sequence":"first","affiliation":[{"name":"Department of Cell Biology and Molecular Genetics, Maryland Pathogen Research Institute University of Maryland  College Park Maryland"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ethan","family":"Iverson","sequence":"additional","affiliation":[{"name":"Department of Cell Biology and Molecular Genetics, Maryland Pathogen Research Institute University of Maryland  College Park Maryland"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Emma J.","family":"DeGrace","sequence":"additional","affiliation":[{"name":"Department of Microbiology Icahn School of Medicine at Mount Sinai  New York New York"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Brad R.","family":"Rosenberg","sequence":"additional","affiliation":[{"name":"Department of Microbiology Icahn School of Medicine at Mount Sinai  New York New York"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Margaret A.","family":"Scull","sequence":"additional","affiliation":[{"name":"Department of Cell Biology and Molecular Genetics, Maryland Pathogen Research Institute University of Maryland  College Park Maryland"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"311","published-online":{"date-parts":[[2022,6,7]]},"reference":[{"key":"e_1_2_8_2_1","doi-asserted-by":"publisher","DOI":"10.1099\/vir.0.031302\u20100"},{"key":"e_1_2_8_3_1","doi-asserted-by":"publisher","DOI":"10.1002\/anbr.202000111"},{"key":"e_1_2_8_4_1","doi-asserted-by":"publisher","DOI":"10.1038\/ng.2830"},{"key":"e_1_2_8_5_1","doi-asserted-by":"publisher","DOI":"10.1186\/s12964\u2010021\u201000740\u2010z"},{"key":"e_1_2_8_6_1","doi-asserted-by":"publisher","DOI":"10.1002\/9780471729259.mc01a01s13"},{"key":"e_1_2_8_7_1","doi-asserted-by":"publisher","DOI":"10.1369\/0022155415570968"},{"key":"e_1_2_8_8_1","doi-asserted-by":"publisher","DOI":"10.1371\/journal.ppat.1010159"},{"key":"e_1_2_8_9_1","first-page":"L347","article-title":"Culture and transformation of human airway epithelial cells","volume":"268","author":"Gruenert D. 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