{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,7]],"date-time":"2026-09-07T22:54:29Z","timestamp":1788821669581,"version":"build-2803163510"},"reference-count":69,"publisher":"MDPI AG","issue":"5","license":[{"start":{"date-parts":[[2026,4,24]],"date-time":"2026-04-24T00:00:00Z","timestamp":1776988800000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/100007482","name":"Arizona State University","doi-asserted-by":"crossref","id":[{"id":"10.13039\/100007482","id-type":"DOI","asserted-by":"crossref"}]},{"id":[{"id":"https:\/\/ror.org\/03efmqc40","id-type":"ROR","asserted-by":"publisher"}]},{"name":"Riney Family Foundation"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Biosensors"],"abstract":"<jats:p>Chimeric antigen receptor (CAR) T-cell therapy is an effective treatment for hematologic malignancies. However, it is limited by high costs, risk of severe toxicities such as cytokine release syndrome and neurotoxicity, and heterogeneous patient responses. The current therapy monitoring depends largely on subjective symptom assessment, routine laboratory tests, and basic vital signs, without real-time, quantitative evaluation of CAR T-cell expansion or activation in clinical practice. This lack of timely immune monitoring hampers individualized care and contributes to increased treatment costs. To address this need, we present a proof-of-concept, label-free rapid optical imaging (ROI) biosensor with automated machine learning analysis for direct quantification of CAR T-cells from whole blood. This microfluidic platform integrates red blood cell (RBC) removal, CAR T-cell capture, and imaging-based quantification on a single chip, eliminating the need for centrifugation, staining, and operator-dependent interpretation. For validation, 50 \u03bcL whole blood samples spiked with Jurkat cells expressing CD19 CARs underwent RBC depletion by agglutination and microfiltration. The remaining blood components were then incubated on a sensor chip functionalized with recombinant CD19 protein. Captured CAR T-cells were imaged by brightfield microscopy and automatically enumerated using a machine learning algorithm trained on fluorescence-validated cells. The CD-19 cells\u2019 capture performance was validated by flow cytometry and fluorescence imaging. The trained machine learning model validated at 88% sensitivity and 96% specificity. Buffer and whole blood calibration curves were established across clinically relevant concentrations (1\u20131000 cells\/\u00b5L) with triple replicates. The results showed high correlation (0.975 and 0.990 R2) between the spiked concentration and the detected CAR T-cells, with a 95% certainty limit of detection (LOD) and quantification (LOQ) of 0.6 and 1.1 cells\/\u00b5L for spiked buffer, and 14 and 67 cells\/\u00b5L for spiked whole-blood, respectively.<\/jats:p>","DOI":"10.3390\/bios16050240","type":"journal-article","created":{"date-parts":[[2026,4,24]],"date-time":"2026-04-24T14:28:23Z","timestamp":1777040903000},"page":"240","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":1,"title":["Machine Learning-Assisted Rapid Optical Imaging for Label-Free CAR T-Cell Detection in Whole Blood"],"prefix":"10.3390","volume":"16","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-8898-4467","authenticated-orcid":false,"given":"Nanxi","family":"Yu","sequence":"first","affiliation":[{"name":"Center for Biosensors and Bioelectronics, The Biodesign Institute, Arizona State University, Tempe, AZ 85287, USA"},{"name":"School of Molecular Sciences, Arizona State University, Tempe, AZ 85287, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0005-4445-2951","authenticated-orcid":false,"given":"Ryan M.","family":"Porter","sequence":"additional","affiliation":[{"name":"Center for Biosensors and Bioelectronics, The Biodesign Institute, Arizona State University, Tempe, AZ 85287, USA"},{"name":"School of Electrical, Computer and Energy Engineering, Arizona State University, Tempe, AZ 85287, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-1399-828X","authenticated-orcid":false,"given":"Xinyu","family":"Zhou","sequence":"additional","affiliation":[{"name":"Center for Biosensors and Bioelectronics, The Biodesign Institute, Arizona State University, Tempe, AZ 85287, USA"},{"name":"School of Biological and Health Systems Engineering, Arizona State University, Tempe, AZ 85287, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-1038-5509","authenticated-orcid":false,"given":"Wenwen","family":"Jing","sequence":"additional","affiliation":[{"name":"Center for Biosensors and Bioelectronics, The Biodesign Institute, Arizona State University, Tempe, AZ 85287, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-4369-8563","authenticated-orcid":false,"given":"Fenni","family":"Zhang","sequence":"additional","affiliation":[{"name":"Center for Biosensors and Bioelectronics, The Biodesign Institute, Arizona State University, Tempe, AZ 85287, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7629-6264","authenticated-orcid":false,"given":"Eider F.","family":"Moreno Cortes","sequence":"additional","affiliation":[{"name":"Division of Hematology & Oncology, Mayo Clinic Arizona, Phoenix, AZ 85054, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-1285-8499","authenticated-orcid":false,"given":"Paula A. Lengerke","family":"Diaz","sequence":"additional","affiliation":[{"name":"Division of Hematology & Oncology, Mayo Clinic Arizona, Phoenix, AZ 85054, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9872-5760","authenticated-orcid":false,"given":"Jose V. Forero","family":"Forero","sequence":"additional","affiliation":[{"name":"Division of Hematology & Oncology, Mayo Clinic Arizona, Phoenix, AZ 85054, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-2407-8251","authenticated-orcid":false,"given":"Erica","family":"Forzani","sequence":"additional","affiliation":[{"name":"Center for Biosensors and Bioelectronics, The Biodesign Institute, Arizona State University, Tempe, AZ 85287, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-2785-0892","authenticated-orcid":false,"given":"Januario E.","family":"Castro","sequence":"additional","affiliation":[{"name":"Division of Hematology & Oncology, Mayo Clinic Arizona, Phoenix, AZ 85054, USA"},{"name":"Division of Hematology & Oncology, Louisiana State University, New Orleans, LA 70112, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-2680-0503","authenticated-orcid":false,"given":"Shaopeng","family":"Wang","sequence":"additional","affiliation":[{"name":"Center for Biosensors and Bioelectronics, The Biodesign Institute, Arizona State University, Tempe, AZ 85287, USA"},{"name":"School of Biological and Health Systems Engineering, Arizona State University, Tempe, AZ 85287, USA"},{"name":"John Shufeldt School of Medicine and Medical Engineering, Arizona State University, Tempe, AZ 85287, USA"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2026,4,24]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"1803","DOI":"10.1200\/JCO.2016.71.3024","article-title":"Lymphoma Remissions Caused by Anti-CD19 Chimeric Antigen Receptor T Cells Are Associated With High Serum Interleukin-15 Levels","volume":"35","author":"Kochenderfer","year":"2017","journal-title":"J. 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