biostudies-literatureArimura YStavros Niarchos FoundationOsamu Hayaishi Memorial ScholarshipNational Institute of General Medical SciencesNIGMS NIH HHSJapan Society for the Promotion of ScienceRP103486https://www.ebi.ac.uk/biostudies/studies/S-EPMC12092007biostudies-literatureUnknown13Cryo-EM single-particle analyses typically require target macromolecule concentration at 0.05~5.0 mg/ml, which is often difficult to achieve. Here, we devise Magnetic Isolation and Concentration (MagIC)-cryo-EM, a technique enabling direct structural analysis of targets captured on magnetic beads, thereby reducing the targets' concentration requirement to <0.0005 mg/mL. Adapting MagIC-cryo-EM to a Chromatin Immunoprecipitation protocol, we characterized structural variations of the linker histone H1.8-associated nucleosomes that were isolated from interphase and metaphase chromosomes in Xenopus egg extract. Combining Duplicated Selection To Exclude Rubbish particles (DuSTER), a particle curation method that excludes low signal-to-noise ratio particles, we also resolved the 3D cryo-EM structures of nucleoplasmin NPM2 co-isolated with the linker histone H1.8 and revealed distinct open and closed structural variants. Our study demonstrates the utility of MagIC-cryo-EM for structural analysis of scarce macromolecules in heterogeneous samples and provides structural insights into the cell cycle-regulation of H1.8 association to nucleosomes.eLifeMagIC-Cryo-EM, structural determination on magnetic beads for scarce macromolecules in heterogeneous samples.PMC12092007SNF Institute for Global Infectious Disease ResearchOverseas Research FellowshipsScholarship for Study AbroadR35 GM132111R35GM132111Funabiki HArimura YKonishi HAfalseMagIC-Cryo-EM, structural determination on magnetic beads for scarce macromolecules in heterogeneous samples.Cryo-EM single-particle analyses typically require target macromolecule concentration at 0.05~5.0 mg/ml, which is often difficult to achieve. Here, we devise Magnetic Isolation and Concentration (MagIC)-cryo-EM, a technique enabling direct structural analysis of targets captured on magnetic beads, thereby reducing the targets' concentration requirement to <0.0005 mg/mL. Adapting MagIC-cryo-EM to a Chromatin Immunoprecipitation protocol, we characterized structural variations of the linker histone H1.8-associated nucleosomes that were isolated from interphase and metaphase chromosomes in Xenopus egg extract. Combining Duplicated Selection To Exclude Rubbish particles (DuSTER), a particle curation method that excludes low signal-to-noise ratio particles, we also resolved the 3D cryo-EM structures of nucleoplasmin NPM2 co-isolated with the linker histone H1.8 and revealed distinct open and closed structural variants. Our study demonstrates the utility of MagIC-cryo-EM for structural analysis of scarce macromolecules in heterogeneous samples and provides structural insights into the cell cycle-regulation of H1.8 association to nucleosomes.2025-01-01T00:00:00Z2025 May2026-06-01T14:56:34.663Z2026-04-08T13:32:32.593ZS-EPMC120920074039036510.7554/eLife.103486