{
  "abstract": "Reporter gene imaging can enable detection of engineered cells in vivo, but tracking hematopoietic engraftment within deep bone marrow (BM) remains challenging. We addressed this challenge by developing a dual CEST-MRI and fluorescent reporter system for cross-validated detection of reporter-positive donor-derived hematopoietic cells and their progeny in BM, with fluorescence-based assessment of HSPC-enriched compartments.Methods\n\nA lentiviral dual-reporter cassette co-expressing the red fluorescent protein mScarlet-I (mScarlet) and the CEST-MRI reporter SuperCESTide (SC) via a T2A linker was evaluated in CT26 colon carcinoma cells in vitro and in a bilateral flank tumor model with contralateral reporter-transduced and non-transduced tumors in BALB/c mice (n = 6 tumors per group). In vivo optical imaging, CEST-MRI, ex vivo optical imaging, fluorescence microscopy, and histology were used for validation. The platform was then applied to primary murine hematopoietic lineage-negative (Lin⁻) cells and Lin⁻Sca-1⁺c-Kit⁺ (LSKs), with reporter expression analyzed by fluorescence microscopy and flow cytometry. Reporter-transduced Lin⁻ cells were transplanted into C57BL/6J mice after half-body irradiation, and engraftment was assessed 8 weeks later by whole-body fluorescence imaging, femoral CEST-MRI, intravital calvarial microscopy, ex vivo confocal microscopy, optical imaging, and flow cytometry. Femoral CEST-MRI was compared among non-irradiated controls, irradiated recipients transplanted with non-reporter-transduced Lin⁻ cells, and irradiated recipients transplanted with mScarlet-SC reporter-transduced Lin⁻ cells.Results\n\nThe dual mScarlet-SuperCESTide reporter showed efficient expression in CT26 cells and generated detectable red fluorescence and increased CEST-MRI contrast in reporter-positive tumors compared with contralateral non-transduced controls. Ex vivo optical imaging and histology confirmed reporter expression in tumor tissue. In murine BM-derived Lin⁻ cells, reporter expression was detected by fluorescence microscopy and flow cytometry, supporting feasibility in sensitive hematopoietic populations. Following transplantation, reporter-transduced Lin⁻ cells were detected eight weeks after engraftment using fluorescence imaging and femoral CEST-MRI. The recipients with reporter-transduced Lin⁻ cells showed increased femoral CEST contrast associated with the reporter compared with both irradiated recipients transplanted with non-transduced Lin⁻ cells and non-irradiated BM controls. Additionally, intravital microscopy visualized mScarlet⁺ cells within the BM niches of the calvarium. Ex vivo confocal microscopy and optical imaging confirmed reporter retention in BM, and flow cytometry confirmed mScarlet signal within Lin⁻ and LSK hematopoietic compartments in reporter-transduced recipients compared with non-transduced controls.Conclusion\n\nWe developed a dual CEST-fluorescent reporter platform that enables cross-validated imaging of reporter-transduced hematopoietic cells in deep BM. By integrating noninvasive CEST-MRI with fluorescence-based cellular validation, this feasibility study establishes a preclinical framework for cross-validated, noninvasive assessment of engineered hematopoietic cell localization and persistence in vivo and supports future evaluation of the platform for serial imaging.",
  "abstract_shown": true,
  "affiliations": [
    "Faculty of Biomedical Engineering, Technion-Israel Institute of Technology, Haifa, Israel.",
    "Biomedical Core Facility, Rappaport Faculty of Medicine, Technion-Israel Institute of Technology, Haifa, Israel.",
    "Biomedical Engineering Faculty, Tel Aviv University, Tel Aviv, Israel.",
    "Chemical Engineering and Radiology, Michigan State University, East Lansing, USA."
  ],
  "article_type": "research article",
  "arxiv": null,
  "author_total": 8,
  "authors": [
    {
      "affs": [
        0
      ],
      "name": "Tehila Alter"
    },
    {
      "affs": [
        0
      ],
      "name": "Narmeen Haj"
    },
    {
      "affs": [
        0
      ],
      "name": "Doaa Ali-Naffaa"
    },
    {
      "affs": [
        0
      ],
      "name": "Maya Rom"
    },
    {
      "affs": [
        0
      ],
      "name": "Ramez Daniel"
    },
    {
      "affs": [
        1
      ],
      "name": "Galit Saar"
    },
    {
      "affs": [
        2,
        3
      ],
      "name": "Assaf A Gilad"
    },
    {
      "affs": [
        0
      ],
      "name": "Katrien Vandoorne"
    }
  ],
  "bibtex": "https://research.draconitic.com/paper/6ctt70q360r0h.bib",
  "cited_by": 0,
  "doi": "10.7150/thno.140697",
  "fulltext_url": "https://europepmc.org/articles/PMC13684844",
  "id": "6ctt70q360r0h",
  "issns": [
    "1838-7640"
  ],
  "landing_url": "https://europepmc.org/article/MED/42909934",
  "language": "eng",
  "licence": "CC-BY",
  "licence_label": "CC BY",
  "licence_source": "article",
  "licence_url": "https://creativecommons.org/licenses/by/4.0/",
  "metadata_licence": null,
  "native_subjects": [
    [
      "openalex",
      "2720|Hematopoietic Stem Cell Transplantation"
    ]
  ],
  "openalex": "W7214677334",
  "pdf_url": "https://europepmc.org/articles/PMC13684844?pdf=render",
  "pmcid": "PMC13684844",
  "pmid": "42909934",
  "published": "2026-09-24",
  "publisher": null,
  "ris": "https://research.draconitic.com/paper/6ctt70q360r0h.ris",
  "sources": [
    {
      "first_seen": "2026-10-30T12:20:14Z",
      "native_id": "MED:42909934",
      "source": "europepmc",
      "url": "https://europepmc.org/article/MED/42909934"
    }
  ],
  "subjects": [
    {
      "field": "medicine",
      "origin": "openalex:2720|Hematopoietic Stem Cell Transplantation",
      "subfield": "general"
    }
  ],
  "title": "A dual CEST-fluorescent reporter enables cross-validated tracking of hematopoietic engraftment in deep bone marrow.",
  "title_text": "A dual CEST-fluorescent reporter enables cross-validated tracking of hematopoietic engraftment in deep bone marrow.",
  "type": "peer-reviewed",
  "updated_at": "2026-10-30T12:20:14Z",
  "url": "https://research.draconitic.com/paper/6ctt70q360r0h",
  "venue": "Theranostics",
  "versions": []
}