Research Center for Applied Sciences, Academia Sinica, Taipei 115201, Taiwan.
Department of Engineering and System Science, National Tsing Hua University, Hsinchu 300044, Taiwan.
Biosensors (Basel). 2024 Feb 11;14(2):96. doi: 10.3390/bios14020096.
Oxygen consumption has been used to evaluate various cellular activities. In addition, three-dimensional (3D) spheroids have been broadly exploited as advanced in vitro cell models for various biomedical studies due to their capability of mimicking 3D in vivo microenvironments and cell arrangements. However, monitoring the oxygen consumption of live 3D spheroids poses challenges because existing invasive methods cause structural and cell damage. In contrast, optical methods using fluorescence labeling and microscopy are non-invasive, but they suffer from technical limitations like high cost, tedious procedures, and poor signal-to-noise ratios. To address these challenges, we developed a microfluidic platform for uniform-sized spheroid formation, handling, and culture. The platform is further integrated with widefield frequency domain fluorescence lifetime imaging microscopy (FD-FLIM) to efficiently characterize the lifetime of an oxygen-sensitive dye filling the platform for oxygen consumption characterization. In the experiments, osteosarcoma (MG-63) cells are exploited as the spheroid model and for the oxygen consumption analysis. The results demonstrate the functionality of the developed approach and show the accurate characterization of the oxygen consumption of the spheroids in response to drug treatments. The developed approach possesses great potential to advance spheroid metabolism studies with single-spheroid resolution and high sensitivity.
氧耗已被用于评估各种细胞活动。此外，由于能够模拟 3D 体内微环境和细胞排列，三维（3D）球体已被广泛用作各种生物医学研究的先进体外细胞模型。然而，监测活 3D 球体的氧耗存在挑战，因为现有的侵入性方法会导致结构和细胞损伤。相比之下，使用荧光标记和显微镜的光学方法是非侵入性的，但它们存在技术限制，如成本高、程序繁琐和信噪比差。为了解决这些挑战，我们开发了一种用于均匀尺寸球体形成、处理和培养的微流控平台。该平台进一步与宽场频域荧光寿命成像显微镜（FD-FLIM）集成，以有效地对填充平台的氧敏染料的寿命进行特征化，从而对氧耗进行特征化。在实验中，骨肉瘤（MG-63）细胞被用作球体模型，并进行氧耗分析。结果证明了所开发方法的功能，并显示了药物处理后球体氧耗的准确特征化。该方法具有以单个球体分辨率和高灵敏度推进球体代谢研究的巨大潜力。
