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Disruption of this ECM in cancer and chronic inflammation has been shown to promote disease progression. While interactions between cells and the LN ECM are critical for immunity, they remain difficult to study due to limitations in current models and reliance on animal studies. To address this, LNs could be decellularized to generate cell-free scaffolds that are subsequently reseeded with cells to study how the native LN microenvironment influences cellular behavior. Existing whole-organ decellularization methods preserve ECM features but yield dense scaffolds that restrict uniform cell seeding, limit nutrient diffusion, and hinder imaging analyses. Here, we present a protocol that combines vibratome sectioning (200-\u03bcm slices) with detergent decellularization (0.1% SDS and 1% Triton-X) to generate thin LN slices from mouse and human tissues. Decellularized LNs had comparable collagen and GAG concentrations to native tissue, and immunofluorescence staining showed the presence of other ECM proteins. Decellularized sections sustained 21-day FRC culture, enabled FRC-T cell co-culture, and supported high-resolution imaging and flow cytometric analyses, revealing altered gp38 and PDGFR\u03b1 expression in FRCs relative to 2D culture.<\/jats:p>","DOI":"10.1038\/s41598-025-28782-0","type":"journal-article","created":{"date-parts":[[2025,11,24]],"date-time":"2025-11-24T10:54:41Z","timestamp":1763981681000},"update-policy":"https:\/\/doi.org\/10.1007\/springer_crossmark_policy","source":"Crossref","is-referenced-by-count":3,"title":["Decellularized lymph node sections with preserved extracellular matrix for stromal cell culture"],"prefix":"10.1038","volume":"16","author":[{"given":"Estefania","family":"Esparza","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Leonor N.","family":"Teles","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Alisa","family":"Fedotova","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Noa","family":"Dehaseth","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Mira","family":"Sayegh","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ana V.","family":"Hernandez","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Lucy Y.","family":"Ho","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Noel M.","family":"Ziebarth","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Alice A.","family":"Tomei","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"297","published-online":{"date-parts":[[2025,11,24]]},"reference":[{"key":"28782_CR1","doi-asserted-by":"publisher","first-page":"136","DOI":"10.1089\/lrb.2013.0012","volume":"11","author":"S Liao","year":"2013","unstructured":"Liao, S. & Padera, T. P. Lymphatic function and immune regulation in health and disease. Lymphat Res. Biol. 11, 136\u2013143. https:\/\/doi.org\/10.1089\/lrb.2013.0012 (2013).","journal-title":"Lymphat Res. Biol."},{"key":"28782_CR2","doi-asserted-by":"publisher","first-page":"723","DOI":"10.1016\/j.it.2021.06.006","volume":"42","author":"L Li","year":"2021","unstructured":"Li, L., Wu, J., Abdi, R., Jewell, C. M. & Bromberg, J. S. Lymph node fibroblastic reticular cells steer immune responses. Trends Immunol. 42, 723\u2013734. https:\/\/doi.org\/10.1016\/j.it.2021.06.006 (2021).","journal-title":"Trends Immunol."},{"key":"28782_CR3","doi-asserted-by":"publisher","first-page":"9","DOI":"10.1111\/imr.12799","volume":"292","author":"V Saxena","year":"2019","unstructured":"Saxena, V. et al. Role of lymph node stroma and microenvironment in T cell tolerance. Immunol. Rev. 292, 9\u201323. https:\/\/doi.org\/10.1111\/imr.12799 (2019).","journal-title":"Immunol. 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