{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,7,10]],"date-time":"2026-07-10T04:15:46Z","timestamp":1783656946762,"version":"3.55.0"},"reference-count":20,"publisher":"The Company of Biologists","issue":"20","license":[{"start":{"date-parts":[[2024,5,28]],"date-time":"2024-05-28T00:00:00Z","timestamp":1716854400000},"content-version":"vor","delay-in-days":0,"URL":"http:\/\/creativecommons.org\/licenses\/by\/4.0"}],"funder":[{"DOI":"10.13039\/100000062","name":"National Institute of Diabetes and Digestive and Kidney Diseases","doi-asserted-by":"crossref","award":["DK115795"],"award-info":[{"award-number":["DK115795"]}],"id":[{"id":"10.13039\/100000062","id-type":"DOI","asserted-by":"crossref"}]},{"DOI":"10.13039\/100000062","name":"National Institute of Diabetes and Digestive and Kidney Diseases","doi-asserted-by":"crossref","award":["DK138974"],"award-info":[{"award-number":["DK138974"]}],"id":[{"id":"10.13039\/100000062","id-type":"DOI","asserted-by":"crossref"}]},{"DOI":"10.13039\/100000062","name":"National Institute of Diabetes and Digestive and Kidney Diseases","doi-asserted-by":"crossref","award":["DK127748"],"award-info":[{"award-number":["DK127748"]}],"id":[{"id":"10.13039\/100000062","id-type":"DOI","asserted-by":"crossref"}]},{"DOI":"10.13039\/100011912","name":"Washington University School of Medicine","doi-asserted-by":"crossref","id":[{"id":"10.13039\/100011912","id-type":"DOI","asserted-by":"crossref"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"published-print":{"date-parts":[[2024,10,15]]},"abstract":"<jats:title>ABSTRACT<\/jats:title>\n               <jats:p>The definitive demonstration of protein localization on primary cilia has been a challenge for cilia biologists. Primary cilia are solitary thread-like projections that have a specialized protein composition, but as the ciliary structure overlays the cell membrane and other cell parts, the identity of ciliary proteins are difficult to ascertain by conventional imaging approaches like immunofluorescence microscopy. Surface scanning electron microscopy combined with immunolabeling (immuno-SEM) bypasses some of these indeterminacies by unambiguously showing protein expression in the context of the three-dimensional ultrastructure of the cilium. Here, we apply immuno-SEM to specifically identify proteins on the primary cilia of mouse and human pancreatic islets, including post-translationally modified tubulin, intraflagellar transport (IFT)88, the small GTPase Arl13b, as well as subunits of axonemal dynein. Key parameters in sample preparation, immunolabeling and imaging acquisition are discussed to facilitate similar studies by others in the cilia research community.<\/jats:p>","DOI":"10.1242\/jcs.262038","type":"journal-article","created":{"date-parts":[[2024,5,28]],"date-time":"2024-05-28T11:41:20Z","timestamp":1716896480000},"source":"Crossref","is-referenced-by-count":11,"title":["Immuno-scanning electron microscopy of islet primary cilia"],"prefix":"10.1242","volume":"137","author":[{"ORCID":"https:\/\/orcid.org\/0000-0001-5918-4834","authenticated-orcid":false,"given":"Sanja","family":"Sviben","sequence":"first","affiliation":[{"name":"Washington University Center for Cellular Imaging, Washington University School of Medicine 1 , 660 South Euclid Ave, Saint Louis, MO 63110 , USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-2601-3638","authenticated-orcid":false,"given":"Alexander J.","family":"Polino","sequence":"additional","affiliation":[{"name":"Washington University School of Medicine 2 Department of Cell Biology and Physiology , , 660 South Euclid Ave, Saint Louis, MO 63110 , USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7894-0682","authenticated-orcid":false,"given":"Isabella L.","family":"Melena","sequence":"additional","affiliation":[{"name":"Washington University School of Medicine 3 Department of Medicine , , 660 South Euclid Ave, Saint Louis, MO 63110 , USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4397-8175","authenticated-orcid":false,"given":"Jing W.","family":"Hughes","sequence":"additional","affiliation":[{"name":"Washington University School of Medicine 2 Department of Cell Biology and Physiology , , 660 South Euclid Ave, Saint Louis, MO 63110 , USA"},{"name":"Washington University School of Medicine 3 Department of Medicine , , 660 South Euclid Ave, Saint Louis, MO 63110 , USA"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"237","published-online":{"date-parts":[[2024,5,28]]},"reference":[{"key":"2024053109154892200_JCS262038C1","doi-asserted-by":"publisher","first-page":"1410","DOI":"10.1073\/pnas.1421845112","article-title":"Intracellular and extracellular forces drive primary cilia movement","volume":"112","author":"Battle","year":"2015","journal-title":"Proc. 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Biol. Clifton NJ"},{"key":"2024053109154892200_JCS262038C5","doi-asserted-by":"publisher","first-page":"e86","DOI":"10.1002\/cpmc.86","article-title":"Immunoelectron microscopy of viral antigens","volume":"53","author":"Gulati","year":"2019","journal-title":"Curr. Protoc. Microbiol."},{"key":"2024053109154892200_JCS262038C6","doi-asserted-by":"publisher","first-page":"5","DOI":"10.1186\/s13630-017-0045-9","article-title":"Fixation methods can differentially affect ciliary protein immunolabeling","volume":"6","author":"Hua","year":"2017","journal-title":"Cilia"},{"key":"2024053109154892200_JCS262038C7","doi-asserted-by":"publisher","first-page":"8912","DOI":"10.1073\/pnas.2001936117","article-title":"Primary cilia control glucose homeostasis via islet paracrine interactions","volume":"117","author":"Hughes","year":"2020","journal-title":"Proc. Natl. Acad. Sci. USA"},{"key":"2024053109154892200_JCS262038C8","doi-asserted-by":"publisher","first-page":"35","DOI":"10.2337\/diab.16.1.35","article-title":"Method for the isolation of intact islets of langerhans from the rat pancreas","volume":"16","author":"Lacy","year":"1967","journal-title":"Diabetes"},{"key":"2024053109154892200_JCS262038C9","doi-asserted-by":"publisher","first-page":"1004136","DOI":"10.3389\/fendo.2022.1004136","article-title":"Fluorescence imaging of beta cell primary cilia","volume":"13","author":"Li","year":"2022","journal-title":"Front. Endocrinol."},{"key":"2024053109154892200_JCS262038C10","doi-asserted-by":"publisher","first-page":"e202010039","DOI":"10.1083\/jcb.202010039","article-title":"3D FIB-SEM reconstruction of microtubule\u2013organelle interaction in whole primary mouse \u03b2 cells","volume":"220","author":"M\u00fcller","year":"2020","journal-title":"J. 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Hoboken NJ"},{"key":"2024053109154892200_JCS262038C14","first-page":"4.26.1","article-title":"Visualization of live primary cilia dynamics using fluorescence microscopy","volume":"Chapter 4","author":"Ott","year":"2012","journal-title":"Curr. Protoc. Cell Biol."},{"key":"2024053109154892200_JCS262038C15","doi-asserted-by":"publisher","first-page":"e2302624120","DOI":"10.1073\/pnas.2302624120","article-title":"Scanning electron microscopy of human islet cilia","volume":"120","author":"Polino","year":"2023","journal-title":"Proc. Natl. Acad. Sci. 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