Iuele Helena, Forciniti Stefania, Onesto Valentina, Colella Francesco, Siciliano Anna Chiara, Chandra Anil, Nobile Concetta, Gigli Giuseppe, Del Mercato Loretta L
Institute of Nanotechnology, National Research Council (CNR-NANOTEC), c/o Campus Ecotekne, via Monteroni, 73100 Lecce, Italy.
Department of Mathematics and Physics ''Ennio De Giorgi", University of Salento, c/o Campus Ecotekne, via Monteroni, 73100 Lecce, Italy.
ACS Appl Mater Interfaces. 2024 Oct 16;16(41):55071-55085. doi: 10.1021/acsami.4c08306. Epub 2024 Aug 28.
Fluorescence imaging allows for noninvasively visualizing and measuring key physiological parameters like pH and dissolved oxygen. In our work, we created two ratiometric fluorescent microsensors designed for accurately tracking dissolved oxygen levels in 3D cell cultures. We developed a simple and cost-effective method to produce hybrid core-shell silica microparticles that are biocompatible and versatile. These sensors incorporate oxygen-sensitive probes (Ru(dpp) or PtOEP) and reference dyes (RBITC or A647 NHS-Ester). SEM analysis confirmed the efficient loading and distribution of the sensing dye on the outer shell. Fluorimetric and CLSM tests demonstrated the sensors' reversibility and high sensitivity to oxygen, even when integrated into 3D scaffolds. Aging and bleaching experiments validated the stability of our hybrid core-shell silica microsensors for 3D monitoring. The Ru(dpp)-RBITC microparticles showed the most promising performance, especially in a pancreatic cancer model using alginate microgels. By employing computational segmentation, we generated 3D oxygen maps during live cell imaging, revealing oxygen gradients in the extracellular matrix and indicating a significant decrease in oxygen level characteristics of solid tumors. Notably, after 12 h, the oxygen concentration dropped to a hypoxic level of PO 2.7 ± 0.1%.
荧光成像能够以非侵入性方式可视化和测量诸如pH值和溶解氧等关键生理参数。在我们的工作中，我们创建了两种比率荧光微传感器，旨在精确跟踪3D细胞培养物中的溶解氧水平。我们开发了一种简单且经济高效的方法来生产具有生物相容性且用途广泛的核壳结构杂化二氧化硅微粒。这些传感器包含氧敏感探针（Ru(dpp)或PtOEP）和参比染料（RBITC或A647 NHS-酯）。扫描电子显微镜（SEM）分析证实了传感染料在外层的有效负载和分布。荧光测定和共聚焦激光扫描显微镜（CLSM）测试表明，即使集成到3D支架中，这些传感器对氧气仍具有可逆性和高灵敏度。老化和漂白实验验证了我们的核壳结构杂化二氧化硅微传感器用于3D监测的稳定性。Ru(dpp)-RBITC微粒表现出最有前景的性能，尤其是在使用藻酸盐微凝胶的胰腺癌模型中。通过采用计算分割技术，我们在活细胞成像过程中生成了3D氧图谱，揭示了细胞外基质中的氧梯度，并表明实体瘤的氧水平特征显著下降。值得注意的是，12小时后，氧浓度降至PO 2.7±0.1%的缺氧水平。
