Training and test images of Glioma C6 cells imaged using phase-contrast microscopy for the task of cell segmentation. The example shows a phase-contrast image of Glioma C6 cells and the manually annotated segmentation mask. Data type: Phase-contrast images with corresponding annotations in COCO format. Microscopy data type: 2D phase-contrast images recorded at 24 or 72-hour intervals after cell seeding. Microscope: BestScope BS-2092 microscope in phase-contrast mode, equipped with 10× and 20× objective lenses. Cell type: C6 glioma cells (rat glial tumor cells, ATCC CCL-107). Image size: 2592 × 1944 px² . File format: .tif (8-bit). File naming convention: The file names include the cultivation time (24h or 72h) and the microscope objective lens (10× or 20×), e.g., spec_24h_10x_17.tif. Dataset subsets: Glioma C6-spec: 45 images captured under strictly controlled imaging conditions, divided into training (30 images), validation (4 images), and test (11 images) subsets. Glioma C6-gen: 30 images captured under varied imaging and seeding conditions, designed to test model generalization. Annotations: Over 20,000 annotated objects across both subsets, including 12,000 cell annotations and 7,800 nucleus annotations. Glioma C6-spec annotations include Type A cells (spheroids formed by cells either unattached to substrates or at the initial stage of cell division), Type B cells (elongated cells with distinct poles, growth phase) and nuclei. Glioma C6-gen subset annotations consist only of general cell instances, without differentiation into types or nuclei. Article reference: "Glioma C6: A Novel Dataset for Training and Benchmarking Cell Segmentation," Malashin et al., 2025. Authors: Roman Malashin¹ ², Svetlana Pashkevich³, Daniil Ilyukhin¹, Arseniy Volkov³, Valeria Yachnaya¹ ², Andrey Denisov³, Maria Mikhalkova¹ Affiliation(s): ¹ Pavlov Institute of Physiology, Russian Academy of Science ² Saint-Petersburg State University of Aerospace Instrumentation, Russia ³ Institute of Physiology, NAS of Belarus