{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"institution":[{"name":"bioRxiv"}],"indexed":{"date-parts":[[2025,12,18]],"date-time":"2025-12-18T21:34:24Z","timestamp":1766093664749,"version":"3.48.0"},"posted":{"date-parts":[[2025,12,16]]},"group-title":"Bioengineering","reference-count":22,"publisher":"openRxiv","license":[{"start":{"date-parts":[[2025,12,16]],"date-time":"2025-12-16T00:00:00Z","timestamp":1765843200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/www.biorxiv.org\/about\/FAQ#license"}],"funder":[{"name":"Stanford Bio-X Interdisciplinary Initiatives Seed Grants Program"},{"name":"Stanford Center for Cancer Nanotechnology Excellence for Translational Diagnostics (CCNE-TD) seed grant"},{"name":"Stanford Beckman Center Technology Development Seed Grants"},{"name":"National Cancer Institute of the National Institutes of Health","award":["R21CA261643"],"award-info":[{"award-number":["R21CA261643"]}]},{"name":"National Cancer Institute of the National Institutes of Health","award":["R01CA283247"],"award-info":[{"award-number":["R01CA283247"]}]},{"name":"Center for Cellular Construction, which is a Science and Technology Center funded by the National Science Foundation","award":["DBI-1548297"],"award-info":[{"award-number":["DBI-1548297"]}]},{"name":"NSF","award":["ECCS-1542152"],"award-info":[{"award-number":["ECCS-1542152"]}]},{"name":"NSF","award":["ECCS-2026822"],"award-info":[{"award-number":["ECCS-2026822"]}]},{"name":"DOE","award":["DE-AC05-76RL01830"],"award-info":[{"award-number":["DE-AC05-76RL01830"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"accepted":{"date-parts":[[2025,12,16]]},"abstract":"<jats:title>ABSTRACT<\/jats:title>\n                <jats:p>To study tissue heterogeneity at the sub-millimeter scale, laser capture microdissection (LCM) has been the leading technology for isolating regions of interest (ROI) for downstream molecular profiling. As the ROI approaches cellular dimensions (\u223c10 \u03bcm), laser-induced photothermal damage and challenges in capturing microtissues in conventional LCM can compromise protein preservation and quantitative fidelity. This work introduces multi-tiered \u03bcDicers, fabricated by two-photon polymerization, to mechanically dissect tissue slices into uniform microtissues down to 10 \u03bcm. The hierarchical blade architecture limits instantaneous blade-tissue engagement and lowers the cutting force relative to single-tier designs. For benchmarking, proteomic analysis is performed on ethanol-fixed human squamous cell carcinoma microtissues generated by \u03bcDicers and by LCM. Under identical Nanodroplet Processing in One pot for Trace Samples (nanoPOTS) and liquid chromatography-mass spectrometry (LC-MS) conditions, \u03bcDicers yield more peptides and proteins than LCM, with the largest gains at 10-20 \u03bcm spatial resolution. Confocal imaging shows catapult-associated cavities in LCM-generated microtissues. This material loss, along with membrane-limited protein extraction, likely reduces protein coverage. In contrast, multi-tiered \u03bcDicers enables reproducible microdissection down to 10 \u03bcm while maintaining high protein coverage. With spatial registration of microtissues under development, \u03bcDicers have potential to complement LCM for next-generation spatial proteomic workflows.<\/jats:p>","DOI":"10.64898\/2025.12.12.694048","type":"posted-content","created":{"date-parts":[[2025,12,16]],"date-time":"2025-12-16T18:10:15Z","timestamp":1765908615000},"source":"Crossref","is-referenced-by-count":0,"title":["A multi-tiered \u03bcDicer with hierarchical blades achieves protein-preserving microdissection down to 10 \u03bcm"],"prefix":"10.64898","author":[{"ORCID":"https:\/\/orcid.org\/0000-0003-3264-4700","authenticated-orcid":false,"given":"Annatoma","family":"Arif","sequence":"first","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Rashmi","family":"Kumar","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Yumi","family":"Kwon","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Ramon","family":"Rodriguez","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Seth C.","family":"Cordts","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Saisneha","family":"Koppaka","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Ying","family":"Zhu","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Ljiljana","family":"Pa\u0161a-Toli\u0107","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Sindy K. Y.","family":"Tang","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"54368","reference":[{"key":"2025121813301130000_2025.12.12.694048v1.1","doi-asserted-by":"publisher","DOI":"10.1038\/s41467-018-03367-w"},{"key":"2025121813301130000_2025.12.12.694048v1.2","doi-asserted-by":"publisher","DOI":"10.1038\/s41587-022-01302-5"},{"key":"2025121813301130000_2025.12.12.694048v1.3","doi-asserted-by":"publisher","DOI":"10.1002\/pmic.202000077"},{"key":"2025121813301130000_2025.12.12.694048v1.4","doi-asserted-by":"publisher","DOI":"10.1038\/s41592-024-02541-x"},{"key":"2025121813301130000_2025.12.12.694048v1.5","doi-asserted-by":"publisher","DOI":"10.1101\/gr.5799207"},{"key":"2025121813301130000_2025.12.12.694048v1.6","doi-asserted-by":"publisher","DOI":"10.1016\/S0168-9525(98)01489-9"},{"key":"2025121813301130000_2025.12.12.694048v1.7","first-page":"1","article-title":"Beyond laser microdissection technology: follow the yellow brick road for cancer research","volume":"4","year":"2014","journal-title":"Am. 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