Live cell imaging of MCF10A cells treated with OSM, IFNG, or EGF, under siRNA knockdown of 14 genes - Replicate 2

This dataset contains live cell imaging of MCF10A mammary epithelial cells subjected to cytokine treatment and siRNA-mediated gene knockdown. MCF10A cells were cultured in DMEM/F12 growth medium supplemented with horse serum, hydrocortisone, cholera toxin, insulin, and EGF, then transitioned to growth factor-free medium prior to treatment. Cells were treated with either Oncostatin M (10 ng/mL), Interferon-gamma plus EGF (IFNG 20 ng/mL + EGF 10 ng/mL), or EGF alone (10 ng/mL). For gene perturbations, siRNA knockdowns targeting 14 candidate regulators were performed using Lipofectamine RNAiMAX-mediated reverse transfection. Knockdown efficency was assessed functionally through phenotypic readouts. Following siRNA transfection and ligand treatment, cells were imaged using the Incucyte S3 microscope for 48 hours, with images acquired every 30 minutes. Fixed-cell immunofluorescent imaging was performed after 48 hours of ligand treatment. Cells were fixed in 4% formaldehyde, permeabilized, and blocked prior to staining with primary conjugated antibodies for B-Catenin and DAPI counterstaining. Imaging was performed using the InCell 6000 platform. This dataset includes raw imaging data cross all conditions (ligand treatments and siRNA knockdowns), enabling the study of cytokine-incuded collective cell migration and phenotypic transitions in mammary epithelial cells. An excel table is included with Well metadata. IF imaging and live cell imaging of the other replicates are available at the linked related Zenodo repositories.本数据集涵盖经细胞因子处理与小干扰RNA（small interfering RNA, siRNA）介导基因敲低的MCF10A乳腺上皮细胞（MCF10A mammary epithelial cells）活细胞成像数据。实验中，MCF10A细胞先于添加马血清、氢化可的松、霍乱毒素、胰岛素及表皮生长因子（Epidermal Growth Factor, EGF）的DMEM/F12完全培养基中培养，随后更换为无生长因子培养基以进行后续处理。细胞分别接受以下三种处理：制瘤素M（Oncostatin M，10 ng/mL）、干扰素γ联合EGF（干扰素γ（IFNG）20 ng/mL + EGF 10 ng/mL），或单独EGF（10 ng/mL）。 针对基因扰动实验，研究人员采用Lipofectamine RNAiMAX介导的反向转染技术，对14个候选调控基因开展siRNA敲低实验。通过表型检测完成敲低效率的功能学评估。在siRNA转染与配体处理完成后，使用Incucyte S3显微镜对细胞进行连续48小时成像，每30分钟采集一次图像。配体处理48小时后，开展固定细胞免疫荧光成像：细胞经4%甲醛固定、透化与封闭处理后，使用针对β-连环蛋白（β-Catenin）的偶联荧光一抗进行染色，并以4',6-二脒基-2-苯基吲哚（DAPI）进行复染，随后通过InCell 6000成像平台完成图像采集。 本数据集包含所有处理条件（配体处理与siRNA敲低组）的原始成像数据，可用于研究细胞因子诱导的乳腺上皮细胞集体迁移与表型转化。数据集附带一份包含孔板元数据的Excel表格。其余重复样本的免疫荧光成像与活细胞成像数据可在相关关联的Zenodo仓储中获取。