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Elucidating their spatiotemporal distribution across biological scales and correlating this distribution with immune activation are of considerable significance. A cross-scale photoacoustic\/fluorescence dual-modality imaging system was developed to enable dynamic tracking of the adjuvant from the cellular to the organismal level. Single-cell imaging results revealed that macrophages exhibited differential activation after the uptake of different adjuvants. Notably, high-resolution photoacoustic microscopy (PAM) revealed size-dependent differences in the intravascular kinetics of adjuvant in zebrafish tail veins as well as their accumulation and clearance during circulation. In a tail amputation-induced inflammation model, fluorescence microscopy demonstrated that CpG oligodeoxynucleotides elicited a 5-fold enhancement in immune cell recruitment efficiency compared to controls, while nano\/micron aluminum adjuvants induced 2-fold and 1.5-fold increases, respectively. The dual-modality photoacoustic\/fluorescence system quantitatively established that these immune outcomes directly originate from the spatiotemporal dynamics of the adjuvant. By bridging single-cell phagocytic behavior with systemic biodistribution, this study deciphered a causal mechanism. The activation of the immune system by adjuvants results from the synergistic interaction of their chemical compositions and spatiotemporal distribution. This imaging-driven framework offers quantitative design principles for precision vaccine adjuvants.<\/jats:p>","DOI":"10.1021\/cbmi.5c00265","type":"journal-article","created":{"date-parts":[[2026,2,13]],"date-time":"2026-02-13T15:22:06Z","timestamp":1770996126000},"page":"1464-1476","source":"Crossref","is-referenced-by-count":0,"title":["Linking Adjuvant Immune\nStimulation to Spatiotemporal\nDynamics via Cross-Scale Photoacoustic Microscopy"],"prefix":"10.1021","volume":"4","author":[{"given":"Dong","family":"Liu","sequence":"first","affiliation":[{"name":"The First Affiliated Hospital of Guangzhou Medical University, Guangzhou Medical University , , ,","place":["Guangzhou, China, 511436"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Fengbing","family":"He","sequence":"additional","affiliation":[{"name":"The First Affiliated Hospital of Guangzhou Medical University, Guangzhou Medical University , , ,","place":["Guangzhou, China, 511436"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yu","family":"Zhang","sequence":"additional","affiliation":[{"name":"The First Affiliated Hospital of Guangzhou Medical University, Guangzhou Medical University , , ,","place":["Guangzhou, China, 511436"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yiqing","family":"Zhang","sequence":"additional","affiliation":[{"name":"The First Affiliated Hospital of Guangzhou Medical University, Guangzhou Medical University , , ,","place":["Guangzhou, China, 511436"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Yibo","family":"Tang","sequence":"additional","affiliation":[{"name":"The First Affiliated Hospital of Guangzhou Medical University, Guangzhou Medical University , , ,","place":["Guangzhou, China, 511436"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Zixi","family":"Liu","sequence":"additional","affiliation":[{"name":"The First Affiliated Hospital of Guangzhou Medical University, Guangzhou Medical University , , ,","place":["Guangzhou, China, 511436"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-0086-4541","authenticated-orcid":true,"given":"Zhangyong","family":"Hong","sequence":"additional","affiliation":[{"name":"Nankai University , , ,","place":["Tianjin, China, 300071"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-9665-9322","authenticated-orcid":true,"given":"Jian","family":"Zhang","sequence":"additional","affiliation":[{"name":"The First Affiliated Hospital of Guangzhou Medical University, Guangzhou Medical University , , ,","place":["Guangzhou, China, 511436"]}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"316","published-online":{"date-parts":[[2026,2,13]]},"reference":[{"issue":"4","key":"2026073101222737300_cit1","doi-asserted-by":"publisher","first-page":"772","DOI":"10.1016\/j.immuni.2024.03.015","article-title":"Vaccine adjuvants: Tailoring innate recognition to\nsend the right message","volume":"57","author":"Lavelle","year":"2024","journal-title":"Immunity"},{"issue":"1","key":"2026073101222737300_cit2","doi-asserted-by":"publisher","first-page":"283","DOI":"10.1038\/s41392-023-01557-7","article-title":"Vaccine adjuvants:\nmechanisms and platforms","volume":"8","author":"Zhao","year":"2023","journal-title":"Signal Transduct Target\nTher."},{"issue":"40","key":"2026073101222737300_cit3","doi-asserted-by":"publisher","first-page":"8166","DOI":"10.1039\/D2TB01358D","article-title":"Polyethyleneimine-based\nimmunoadjuvants\nfor designing cancer vaccines","volume":"10","author":"Yang","year":"2022","journal-title":"J. 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