{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,28]],"date-time":"2026-09-28T12:58:59Z","timestamp":1790600339401,"version":"4.1.0"},"reference-count":51,"publisher":"American Chemical Society (ACS)","issue":"9","license":[{"start":{"date-parts":[[2026,3,27]],"date-time":"2026-03-27T00:00:00Z","timestamp":1774569600000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/100000052","name":"NIH Office of the Director","doi-asserted-by":"publisher","award":["1R35GM157103"],"award-info":[{"award-number":["1R35GM157103"]}],"id":[{"id":"10.13039\/100000052","id-type":"DOI","asserted-by":"publisher"}]},{"DOI":"10.13039\/100000078","name":"Division of Materials Research","doi-asserted-by":"publisher","award":["DMR-1420073"],"award-info":[{"award-number":["DMR-1420073"]}],"id":[{"id":"10.13039\/100000078","id-type":"DOI","asserted-by":"publisher"}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":[],"published-print":{"date-parts":[[2026,9,28]]},"abstract":"<jats:title>Abstract<\/jats:title>\n                  <jats:p>Biomolecular condensates formed through phase separation are fundamental to the cellular organization. Although the physical principles underlying the phase separation of intrinsically disordered proteins are well understood, the molecular determinants of condensate formation in globular proteins remain elusive. Here, we employ holographic particle characterization, a label-free, high-throughput imaging technique, to investigate the self-assembly of bovine serum albumin (BSA), a model globular protein. We show that this technique reliably differentiates between liquid-like and gel-like condensates by their distinct refractive indices and morphologies. Coupled with size-exclusion chromatography, our analysis reveals that BSA phase separation strictly depends on higher-order oligomeric assemblies. Monomeric and dimeric fractions failed to form condensates under identical crowding conditions. Furthermore, the internal packing density of these condensates is tunable via pH-driven protonation changes but remains insensitive to ionic screening. These findings support a model of \u201cemergent multivalency,\u201d in which oligomerization increases the effective interaction valence and creates a structural scaffold that enables hydrophobically stabilized phase separation, consistent with phase separation coupled to percolation frameworks.<\/jats:p>","DOI":"10.1021\/cbmi.5c00258","type":"journal-article","created":{"date-parts":[[2026,3,27]],"date-time":"2026-03-27T11:21:46Z","timestamp":1774610506000},"page":"2023-2031","source":"Crossref","is-referenced-by-count":2,"title":["Holographic Fingerprinting\nReveals Oligomer-Driven\nPhase Separation in Bovine Serum Albumin"],"prefix":"10.1021","volume":"4","author":[{"ORCID":"https:\/\/orcid.org\/0000-0003-3007-3350","authenticated-orcid":true,"given":"Fook Chiong","family":"Cheong","sequence":"first","affiliation":[{"name":"New York University , , , ,","place":["New York, New York, United States, 10003"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0001-7864-9462","authenticated-orcid":true,"given":"Seung Y.","family":"Lee","sequence":"additional","affiliation":[{"name":"New York University , , , ,","place":["New York, New York, United States, 10003"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Sujata","family":"Bais","sequence":"additional","affiliation":[{"name":"New York University , , , ,","place":["New York, New York, United States, 10003"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Satyam","family":"Khanal","sequence":"additional","affiliation":[{"name":"New York University , , , ,","place":["New York, New York, United States, 10003"]}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-7524-7548","authenticated-orcid":true,"given":"Saumya","family":"Saurabh","sequence":"additional","affiliation":[{"name":"New York University , , , ,","place":["New York, New York, United States, 10003"]}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"316","published-online":{"date-parts":[[2026,3,27]]},"reference":[{"key":"2026092808070109100_cit1","doi-asserted-by":"publisher","first-page":"285","DOI":"10.1038\/nrm.2017.7","article-title":"Biomolecular condensates:\norganizers of Cellular Biochemistry","volume":"18","author":"Banani","year":"2017","journal-title":"Nat. Rev.\nMol. Cell Biol."},{"key":"2026092808070109100_cit2","doi-asserted-by":"publisher","first-page":"215","DOI":"10.1038\/s41580-020-00303-z","article-title":"A framework\nfor understanding the\nfunctions of biomolecular condensates across scales","volume":"22","author":"Lyon","year":"2021","journal-title":"Nat. Rev. Mol. Cell. Biol."},{"key":"2026092808070109100_cit3","doi-asserted-by":"publisher","first-page":"2201","DOI":"10.1016\/j.molcel.2022.05.018","article-title":"A conceptual framework\nfor understanding phase separation\nand addressing open questions and challenges","volume":"82","author":"Mittag","year":"2022","journal-title":"Mol. Cell"},{"key":"2026092808070109100_cit4","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.2017184118","article-title":"Ligand effects on phase separation of multivalent macromolecules","volume":"118","author":"Ruff","year":"2021","journal-title":"Proc. Natl. Acad. Sci. U. S. A."},{"key":"2026092808070109100_cit5","doi-asserted-by":"publisher","first-page":"107","DOI":"10.1146\/annurev-biophys-121219-081629","article-title":"Physical principles underlying the\ncomplex biology of intracellular phase transitions","volume":"49","author":"Choi","year":"2020","journal-title":"Annual review of biophysics"},{"key":"2026092808070109100_cit6","doi-asserted-by":"publisher","first-page":"5908","DOI":"10.3390\/ijms21165908","article-title":"Liquid\u2013liquid\nphase separation in crowded environments","volume":"21","author":"Andre\u0301","year":"2020","journal-title":"International\nJournal of Molecular Sciences"},{"key":"2026092808070109100_cit7","doi-asserted-by":"publisher","DOI":"10.1016\/j.abb.2025.110620","article-title":"Temperature and cosolute\nregulate the liquid-liquid phase separation in BSA solutions","volume":"774","author":"Naranjo","year":"2025","journal-title":"Arch. Biochem. Biophys."},{"key":"2026092808070109100_cit8","doi-asserted-by":"publisher","first-page":"3636","DOI":"10.1021\/acs.jpclett.2c00307","article-title":"Macromolecular\ncrowding-induced unusual liquid\u2013liquid phase separation of\nhuman serum albumin via soft protein\u2013protein interactions","volume":"13","author":"Patel","year":"2022","journal-title":"J. Phys. Chem. Lett."},{"key":"2026092808070109100_cit9","doi-asserted-by":"publisher","first-page":"2060","DOI":"10.1021\/acs.biomac.4c00609","article-title":"Protein-Specific\nCrowding Accelerates Aging in Protein Condensates","volume":"26","author":"Brzezinski","year":"2025","journal-title":"Biomacromolecules"},{"key":"2026092808070109100_cit10","doi-asserted-by":"publisher","first-page":"42","DOI":"10.1016\/j.ijbiomac.2012.10.030","article-title":"Bovine serum albumin oligomers in\nthe E-and B-forms\nat low protein concentration and ionic strength","volume":"53","author":"Babcock","year":"2013","journal-title":"Int. J. Biol. Macromol."},{"key":"2026092808070109100_cit11","doi-asserted-by":"publisher","first-page":"430","DOI":"10.1080\/03639045.2025.2479758","article-title":"Formation of self-assembled\npolyelectrolyte complex derived from BSA and nanogels: a study to\noptimize processing parameters and preserve protein integrity","volume":"51","author":"Mudassir","year":"2025","journal-title":"Drug Dev. Ind. Pharm."},{"key":"2026092808070109100_cit12","doi-asserted-by":"publisher","DOI":"10.1016\/j.molliq.2021.118183","article-title":"On the\naggregation of bovine serum albumin","volume":"349","author":"Madeira","year":"2022","journal-title":"J. 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Colloid Interface Sci."},{"key":"2026092808070109100_cit15","doi-asserted-by":"publisher","first-page":"10738","DOI":"10.1021\/acsnano.4c01534","article-title":"Label-free techniques for probing\nbiomolecular condensates","volume":"18","author":"Ibrahim","year":"2024","journal-title":"ACS Nano"},{"key":"2026092808070109100_cit16","doi-asserted-by":"publisher","first-page":"13071","DOI":"10.1364\/OE.17.013071","article-title":"Flow visualization\nand flow cytometry with holographic\nvideo microscopy","volume":"17","author":"Cheong","year":"2009","journal-title":"Opt. 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Pharm. Sci."},{"key":"2026092808070109100_cit22","doi-asserted-by":"publisher","first-page":"25242","DOI":"10.1021\/jacs.5c02947","article-title":"Multivalency\ncontrols the growth and dynamics of a biomolecular condensate","volume":"147","author":"von Hofe","year":"2025","journal-title":"J. Am. Chem. Soc."},{"key":"2026092808070109100_cit23","doi-asserted-by":"publisher","first-page":"336","DOI":"10.1038\/nature10879","article-title":"Phase transitions in the assembly of multivalent signalling proteins","volume":"483","author":"Li","year":"2012","journal-title":"Nature"},{"key":"2026092808070109100_cit24","doi-asserted-by":"publisher","first-page":"1772","DOI":"10.1021\/acs.macromol.8b02476","article-title":"Impact of macromolecular\ncrowding and compression on protein\u2013protein interactions and\nliquid\u2013liquid phase separation phenomena","volume":"52","author":"Julius","year":"2019","journal-title":"Macromolecules"},{"key":"2026092808070109100_cit25","doi-asserted-by":"publisher","first-page":"13480","DOI":"10.1073\/pnas.1921617117","article-title":"Depletion\ninteractions modulate the binding between disordered proteins in crowded\nenvironments","volume":"117","author":"Zosel","year":"2020","journal-title":"Proc. Natl. Acad. Sci. U. S. 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Chem."},{"key":"2026092808070109100_cit31","doi-asserted-by":"publisher","DOI":"10.7554\/eLife.04123","article-title":"Phase transitions\nof multivalent proteins can promote\nclustering of membrane receptors","volume":"3","author":"Banjade","year":"2014","journal-title":"eLife"},{"key":"2026092808070109100_cit32","doi-asserted-by":"publisher","first-page":"2025.06.23.660947","DOI":"10.1101\/2025.06.23.660947","article-title":"Phase separation\nof PGL-3 driven by structured domains that oligomerize and interact\nwith terminal RGG motifs","author":"Kuroiwa","year":"2025","journal-title":"bioRxiv"},{"key":"2026092808070109100_cit33","doi-asserted-by":"publisher","first-page":"87","DOI":"10.1093\/jb\/mvaf065","article-title":"Coiled-coils as emerging drivers\nof liquid\u2013liquid phase separation","volume":"179","author":"Anzai","year":"2026","journal-title":"J.\nBiochem."},{"key":"2026092808070109100_cit34","doi-asserted-by":"publisher","DOI":"10.1016\/j.jmb.2020.166731","article-title":"Liquid-liquid phase\nseparation of tau driven by hydrophobic interaction facilitates fibrillization\nof tau","volume":"433","author":"Lin","year":"2021","journal-title":"Journal of molecular biology"},{"key":"2026092808070109100_cit35","doi-asserted-by":"publisher","first-page":"28a","DOI":"10.1016\/j.bpj.2020.11.426","article-title":"Reentrant\nliquid condensate phase of proteins is stabilized by hydrophobic and\nnon-ionic interactions","volume":"120","author":"Krainer","year":"2021","journal-title":"Biophys. J."},{"key":"2026092808070109100_cit36","doi-asserted-by":"publisher","first-page":"3383","DOI":"10.1021\/jacs.3c12789","article-title":"Surface charge can modulate phase\nseparation of multidomain proteins","volume":"146","author":"Kim","year":"2024","journal-title":"J. Am. Chem.\nSoc."},{"key":"2026092808070109100_cit37","doi-asserted-by":"publisher","DOI":"10.1002\/adom.202100697","article-title":"Label-Free Quantitative Analysis\nof Coacervates via 3D Phase Imaging","volume":"9","author":"Hong","year":"2021","journal-title":"Adv. Opt.\nMater."},{"key":"2026092808070109100_cit38","doi-asserted-by":"publisher","first-page":"372","DOI":"10.1016\/j.tibtech.2014.05.005","article-title":"Therapeutic\nprotein aggregation: mechanisms, design, and control","volume":"32","author":"Roberts","year":"2014","journal-title":"Trends Biotechnol."},{"key":"2026092808070109100_cit39","doi-asserted-by":"publisher","first-page":"10568","DOI":"10.3390\/ijms262110568","article-title":"Current Understanding\nof Protein\nAggregation in Neurodegenerative Diseases","volume":"26","author":"Hu","year":"2025","journal-title":"International\nJournal of Molecular Sciences"},{"key":"2026092808070109100_cit40","doi-asserted-by":"publisher","first-page":"1075","DOI":"10.3724\/abbs.2023106","article-title":"Small molecules in regulating protein\nphase separation: Regulating protein phase separation by small molecules","volume":"55","author":"Li","year":"2023","journal-title":"Acta biochimica et biophysica Sinica"},{"key":"2026092808070109100_cit41","doi-asserted-by":"publisher","DOI":"10.1096\/fj.202501476R","article-title":"Small Molecules\nas Regulators of Liquid\u2013Liquid Phase Separation: Mechanisms\nand Strategies for New Drug Discovery","volume":"39","author":"Usman","year":"2025","journal-title":"FASEB\nJ."},{"key":"2026092808070109100_cit42","doi-asserted-by":"publisher","first-page":"25692","DOI":"10.1021\/jacs.5c07295","article-title":"Selective ion binding and uptake shape the microenvironment\nof biomolecular condensates","volume":"147","author":"Smokers","year":"2025","journal-title":"J. Am. Chem. Soc."},{"key":"2026092808070109100_cit43","doi-asserted-by":"publisher","first-page":"694","DOI":"10.1126\/science.aaw8653","article-title":"Valence and\npatterning of aromatic residues determine\nthe phase behavior of prion-like domains","volume":"367","author":"Martin","year":"2020","journal-title":"Science"},{"key":"2026092808070109100_cit44","doi-asserted-by":"publisher","DOI":"10.1126\/sciadv.abm6570","article-title":"ATP-responsive\nbiomolecular condensates\ntune bacterial kinase signaling","volume":"8","author":"Saurabh","year":"2022","journal-title":"Sci. Adv."},{"key":"2026092808070109100_cit45","doi-asserted-by":"publisher","DOI":"10.1371\/journal.ppat.1012413","article-title":"Biomolecular\ncondensates\nas stress sensors and modulators of bacterial signaling","volume":"20","author":"Sasazawa","year":"2024","journal-title":"PLoS Pathog."},{"key":"2026092808070109100_cit46","doi-asserted-by":"publisher","first-page":"180","DOI":"10.1016\/j.sbi.2016.08.001","article-title":"Liquid\u2013liquid phase separation\nin cellular signaling\nsystems","volume":"41","author":"Chong","year":"2016","journal-title":"Curr. Opin. Struct. Biol."},{"key":"2026092808070109100_cit47","doi-asserted-by":"publisher","first-page":"676","DOI":"10.1038\/nmeth.2019","article-title":"Fiji: an open-source\nplatform for biological-image analysis","volume":"9","author":"Schindelin","year":"2012","journal-title":"Nat.\nMethods"},{"key":"2026092808070109100_cit48","doi-asserted-by":"publisher","first-page":"18275","DOI":"10.1364\/OE.15.018275","article-title":"Characterizing and\ntracking single colloidal particles with video holographic microscopy","volume":"15","author":"Lee","year":"2007","journal-title":"Opt. Express"},{"key":"2026092808070109100_cit49","doi-asserted-by":"publisher","first-page":"83","DOI":"10.1038\/s43586-022-00165-z","article-title":"In-line\nholographic microscopy with model-based analysis","volume":"2","author":"Martin","year":"2022","journal-title":"Nat. Rev. Methods Primers"},{"key":"2026092808070109100_cit50","doi-asserted-by":"publisher","first-page":"494","DOI":"10.1037\/0033-2909.114.3.494","article-title":"Dominance\nstatistics: Ordinal analyses to answer ordinal questions","volume":"114","author":"Cliff","year":"1993","journal-title":"Psychol. Bull."},{"key":"2026092808070109100_cit51","first-page":"1","article-title":"Exploring\nmethods\nfor evaluating group differences on the NSSE and other surveys: Are\nthe t-test and Cohen\u2019s d indices the most appropriate choices?","author":"Romano","year":"2006","journal-title":"Annu. Meet. South. Assoc. Inst. Res."}],"container-title":["Chemical &amp; Biomedical\nImaging"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/pubs.acs.org\/doi\/pdf\/10.1021\/cbmi.5c00258","content-type":"application\/pdf","content-version":"vor","intended-application":"unspecified"},{"URL":"https:\/\/pubs.acs.org\/cbihbp\/article-pdf\/4\/9\/2023\/66262161\/cbmi.5c00258.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"syndication"},{"URL":"https:\/\/pubs.acs.org\/cbihbp\/article-pdf\/4\/9\/2023\/66262161\/cbmi.5c00258.pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2026,9,28]],"date-time":"2026-09-28T12:09:38Z","timestamp":1790597378000},"score":1,"resource":{"primary":{"URL":"https:\/\/pubs.acs.org\/cbihbp\/article\/4\/9\/2023\/5232863\/Holographic-Fingerprinting-Reveals-Oligomer-Driven"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2026,3,27]]},"references-count":51,"journal-issue":{"issue":"9","published-online":{"date-parts":[[2026,3,27]]},"published-print":{"date-parts":[[2026,9,28]]}},"URL":"https:\/\/doi.org\/10.1021\/cbmi.5c00258","relation":{},"ISSN":["2832-3637"],"issn-type":[{"value":"2832-3637","type":"electronic"}],"subject":[],"published":{"date-parts":[[2026,3,27]]}}}