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Although the physical principles underlying 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 amorphous aggregates from liquid-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 fail to form condensates under identical crowding conditions. Furthermore, the internal packing density of these condensates is highly tunable via pH-driven protonation changes but remains insensitive to ionic screening. These findings support a model of \u201cemergent multivalency,\u201d where oligomerization creates a structural scaffold enabling hydrophobically stabilized phase separation, thereby defining a molecular threshold for globular proteins condensation.<\/jats:p>","DOI":"10.64898\/2025.12.10.693513","type":"posted-content","created":{"date-parts":[[2025,12,13]],"date-time":"2025-12-13T22:55:11Z","timestamp":1765666511000},"source":"Crossref","is-referenced-by-count":0,"title":["Holographic fingerprinting reveals oligomer-driven phase separation in Bovine Serum Albumin"],"prefix":"10.64898","author":[{"given":"Fook C.","family":"Cheong","sequence":"first","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Seung Y.","family":"Lee","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Sujata","family":"Bais","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Satyam","family":"Khanal","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Saumya","family":"Saurabh","sequence":"additional","affiliation":[],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"54368","reference":[{"key":"2025121613301091000_2025.12.10.693513v1.1","doi-asserted-by":"publisher","DOI":"10.1038\/nrm.2017.7"},{"key":"2025121613301091000_2025.12.10.693513v1.2","doi-asserted-by":"publisher","DOI":"10.1038\/s41580-020-00303-z"},{"key":"2025121613301091000_2025.12.10.693513v1.3","doi-asserted-by":"publisher","DOI":"10.1016\/j.molcel.2022.05.018"},{"key":"2025121613301091000_2025.12.10.693513v1.4","doi-asserted-by":"publisher","DOI":"10.1073\/pnas.2017184118"},{"key":"2025121613301091000_2025.12.10.693513v1.5","doi-asserted-by":"publisher","DOI":"10.1146\/annurev-biophys-121219-081629"},{"key":"2025121613301091000_2025.12.10.693513v1.6","doi-asserted-by":"crossref","first-page":"196","DOI":"10.1038\/s41557-021-00840-w","article-title":"Deciphering how naturally occurring sequence features impact the phase behaviours of disordered prion-like domains","volume":"14","year":"2022","journal-title":"Nature chemistry"},{"key":"2025121613301091000_2025.12.10.693513v1.7","doi-asserted-by":"crossref","first-page":"5908","DOI":"10.3390\/ijms21165908","article-title":"Liquid\u2013liquid phase separation in crowded environments","volume":"21","year":"2020","journal-title":"International Journal of Molecular Sciences"},{"key":"2025121613301091000_2025.12.10.693513v1.8","doi-asserted-by":"crossref","unstructured":"Naranjo, B. 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