We present a method for real‐time visualization and automatic processing for detection and classification of untreated cancer cells in blood during stain‐free imaging flow cytometry using digital holographic microscopy and machine learning in throughput of 15 cells per second. As a preliminary model for circulating tumor cells in the blood, following an initial label‐free rapid enrichment stage based on the cell size, we applied our holographic imaging approach, providing the quantitative optical thickness profiles of the cells during flow. We automatically classified primary and metastatic colon cancer cells, where the two types of cancer cells were isolated from the same individual, as well as four types of blood cells. We used low‐coherence off‐axis interferometric phase microscopy and a microfluidic channel to image cells during flow quantitatively. The acquired images were processed and classified based on their morphology and quantitative phase features during the cell flow. We achieved high accuracy of 92.56% for distinguishing between the cells, enabling further automatic enrichment and cancer‐cell grading from blood. © 2020 International Society for Advancement of Cytometry

We present a method for label-free imaging and sorting of cancer cells in blood, which is based on a dielectrophoretic microfluidic chip and labelfree interferometric phase microscopy. The chip used for imaging has been embedded with dielectrophoretic electrodes, and therefore it can be used to sort the cells based on the decisions obtained during the cell flow by the label-free quantitative imaging method. Hence, we obtained a real-time, automatic, label-free imaging flow cytometry with the ability to sort the cells during flow. To validate our model, we combined into the label-free imaging interferometer a fluorescence imaging channel that indicated the correctness of the label-free sorting. We have achieved above 98% classification success and 69% sorting accuracy at flow rates of 4 to 7 μL hr −1. In the future, this method is expected to help in label-free sorting of circulating tumor cells in blood following an initial state-of-the-art cell enrichment.

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