{
  "abstract": "Binding affinity measurements are essential for optimizing and validating therapeutic and research antibodies, yet no high-throughput platform supports quantitative affinity measurements for both soluble and native membrane targets. Here we present a suite of digital flow cytometry (dFC) assays that determine equilibrium binding affinities by directly counting free and bound molecules in homogeneous mixtures without immobilization. The assays enable titration-free absolute KD measurements for fluorescent binding partners, affinity measurements for unlabeled antibodies against membrane receptors on cell-derived extracellular vesicles (EVs), and multiplexed assays for rapid ranking or screening of antibody variants. We demonstrate applications by quantifying the effects of degree-of-labeling on the affinity of antibody-dye conjugates, and of drug conjugation on therapeutic antibody binding to receptors on human cancer EVs. By combining single-molecule quantitation, adaptable assay designs, and high-throughput analysis, dFC provides a versatile platform for therapeutic antibody optimization and research antibody validation.",
  "abstract_shown": true,
  "affiliations": [
    "University of Washington"
  ],
  "article_type": "New results",
  "arxiv": null,
  "author_total": 6,
  "authors": [
    {
      "name": "Y. Cheng"
    },
    {
      "name": "S. Barcy"
    },
    {
      "name": "B. Fujimoto"
    },
    {
      "name": "W. Nelson"
    },
    {
      "name": "M. Zhao"
    },
    {
      "affs": [
        0
      ],
      "name": "D. T. Chiu"
    }
  ],
  "bibtex": "https://research.draconitic.com/paper/2179anrk5ed9r.bib",
  "cited_by": 0,
  "doi": "10.64898/2026.10.15.759441",
  "fulltext_url": "https://www.biorxiv.org/content/10.64898/2026.10.15.759441v1.full",
  "id": "2179anrk5ed9r",
  "issns": [
    "2692-8205"
  ],
  "landing_url": "https://www.biorxiv.org/content/10.64898/2026.10.15.759441v1",
  "language": "en",
  "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",
      "biophysics"
    ],
    [
      "heading",
      "Article"
    ],
    [
      "openalex",
      "2741|Monoclonal and Polyclonal Antibodies Research"
    ]
  ],
  "openalex": "W7222517583",
  "pdf_url": "https://www.biorxiv.org/content/10.64898/2026.10.15.759441v1.full.pdf",
  "pmcid": "PMC13738427",
  "pmid": "42995821",
  "published": "2026-10-15",
  "publisher": "Cold Spring Harbor Laboratory Preprints",
  "ris": "https://research.draconitic.com/paper/2179anrk5ed9r.ris",
  "sources": [
    {
      "first_seen": "2026-10-30T12:28:05Z",
      "native_id": "10.64898/2026.10.15.759441",
      "source": "biorxiv",
      "url": "https://www.biorxiv.org/content/10.64898/2026.10.15.759441v2"
    },
    {
      "first_seen": "2026-11-09T12:23:24Z",
      "native_id": "PMC13738427",
      "source": "pmc",
      "url": "https://doi.org/10.64898/2026.10.15.759441"
    }
  ],
  "subjects": [
    {
      "field": "biochemistry",
      "origin": "biorxiv:biophysics",
      "subfield": "biophysics"
    }
  ],
  "title": "High-throughput affinity profiling of antibodies against soluble and native membrane targets by digital flow cytometry",
  "title_text": "High-throughput affinity profiling of antibodies against soluble and native membrane targets by digital flow cytometry",
  "type": "preprint",
  "updated_at": "2026-11-09T12:23:24Z",
  "url": "https://research.draconitic.com/paper/2179anrk5ed9r",
  "venue": "bioRxiv",
  "versions": []
}