{
  "abstract": "Multiplexed imaging technologies are transforming the study of human tissue biology, but their application to the aged brain is hindered by autofluorescence, particularly in fresh-frozen specimens. Here, we characterized autofluorescence across four brain regions from 21 donors and found broad spectral emission, regional and gray-white matter differences, and an association with donor age. Photobleaching conditions adopted from formalin-fixed paraffin-embedded tissue caused marked region- and compartment-dependent damage in fresh-frozen sections. We therefore developed a Tris-EDTA-supplemented photobleaching workflow that reduced autofluorescence by 58-70% while preserving tissue architecture and cellular content. We established a custom 28-plex DNA-barcoded antibody panel targeting neuronal, glial, immune, and vascular markers, providing a resource for fresh-frozen human brain. Integration of the optimized photobleaching workflow with this panel enabled spatial proteomic analysis across fresh-frozen brain regions. By co-registering pre-photobleaching autofluorescence with multiplexed protein maps, we further established a cellular-resolution framework for spatial characterization of autofluorescence. This revealed region-dependent protein marker relationships and preferential enrichment of autofluorescent particles near nuclei and within microglial and CD68-positive regions. Together, this work establishes a practical workflow for multiplexed spatial proteomics in fresh-frozen brain and characterizes autofluorescence as both a technical confound and a spatially structured feature of the aged human brain.",
  "abstract_shown": true,
  "affiliations": [
    "KTH Royal Institute of Technology - SciLifeLab"
  ],
  "article_type": "New results",
  "arxiv": null,
  "author_total": 6,
  "authors": [
    {
      "name": "I. Sutevski"
    },
    {
      "name": "L. Guo"
    },
    {
      "name": "N. Branzell"
    },
    {
      "name": "P. E. Andren"
    },
    {
      "name": "P. Svenningsson"
    },
    {
      "affs": [
        0
      ],
      "name": "B. Ayoglu"
    }
  ],
  "bibtex": "https://research.draconitic.com/paper/67rma96x51k6n.bib",
  "cited_by": 0,
  "doi": "10.64898/2026.09.11.750056",
  "fulltext_url": "https://www.biorxiv.org/content/10.64898/2026.09.11.750056v1.full",
  "id": "67rma96x51k6n",
  "issns": [],
  "landing_url": "https://www.biorxiv.org/content/10.64898/2026.09.11.750056v1",
  "language": null,
  "licence": "CC-BY-NC-ND",
  "licence_label": "CC BY-NC-ND",
  "licence_source": "article",
  "licence_url": "https://creativecommons.org/licenses/by-nc-nd/4.0/",
  "metadata_licence": null,
  "native_subjects": [
    [
      "biorxiv",
      "neuroscience"
    ],
    [
      "openalex",
      "1312|Single-cell and spatial transcriptomics"
    ]
  ],
  "openalex": "W7213552502",
  "pdf_url": "https://www.biorxiv.org/content/10.64898/2026.09.11.750056v1.full.pdf",
  "pmcid": null,
  "pmid": null,
  "published": "2026-09-11",
  "publisher": null,
  "ris": "https://research.draconitic.com/paper/67rma96x51k6n.ris",
  "sources": [
    {
      "first_seen": "2026-10-30T12:23:06Z",
      "native_id": "10.64898/2026.09.11.750056",
      "source": "biorxiv",
      "url": "https://www.biorxiv.org/content/10.64898/2026.09.11.750056v1"
    }
  ],
  "subjects": [
    {
      "field": "biology",
      "origin": "biorxiv:neuroscience",
      "subfield": "neuroscience"
    }
  ],
  "title": "Spatial organization and mitigation of autofluorescence in multiplexed spatial proteomics of aged fresh-frozen human brain",
  "title_text": "Spatial organization and mitigation of autofluorescence in multiplexed spatial proteomics of aged fresh-frozen human brain",
  "type": "preprint",
  "updated_at": "2026-10-30T12:23:06Z",
  "url": "https://research.draconitic.com/paper/67rma96x51k6n",
  "venue": "bioRxiv",
  "versions": []
}