追踪整个群体中的单个细胞通路对于理解他们的行为至关重要,细胞通讯,迁移动态,和命运。光学标记是追踪单个细胞的一种方法,但它通常需要遗传修饰来诱导光转化蛋白的产生。然而,这种方法有局限性,不适用于某些细胞类型。例如,遗传修饰常导致巨噬细胞死亡。这项研究旨在通过利用光可转换的微米大小的胶囊来开发一种标记巨噬细胞的替代方法,该胶囊能够容易内化并延长细胞内的滞留时间。在聚乙烯醇凝胶介质中的热处理用于具有多种荧光染料的胶囊的可扩展合成,包括罗丹明6G,维罗宁B,荧光素,吖啶黄,吖啶橙,噻嗪红,以前报道过罗丹明B的荧光亮度,光稳定性,使用共聚焦激光扫描显微镜评估胶囊的光转换能力。活力,摄取,移动性,在RAW264.7和骨髓来源的巨噬细胞上进行光转换研究,作为模型细胞系。由于使用聚乙烯醇凝胶,胶囊的产量增加,消除了对常规过滤步骤的需要。包埋罗丹明B和罗丹明6G的胶囊满足在单个细胞追踪中细胞内使用的所有要求。质谱分析揭示了一系列脱乙基步骤,这些步骤在辐照时导致染料光谱中的蓝移。对巨噬细胞的细胞研究表明胶囊的强烈摄取。胶囊表现出最小的细胞毒性,对细胞运动性的影响可忽略不计。细胞内含有RhB的胶囊的成功光转换凸显了它们作为单个细胞标记的光转换蛋白的替代品的潜力。在个性化医疗中具有广阔的应用前景。
Tracing individual cell pathways among the whole population is crucial for understanding their behavior, cell communication, migration dynamics, and fate. Optical labeling is one approach for tracing individual cells, but it typically requires genetic modification to induce the generation of photoconvertible proteins. Nevertheless, this approach has limitations and is not applicable to certain cell types. For instance, genetic modification often leads to the death of macrophages. This study aims to develop an alternative method for labeling macrophages by utilizing photoconvertible micron-sized capsules capable of easy internalization and prolonged retention within cells. Thermal treatment in a polyvinyl alcohol gel medium is employed for the scalable synthesis of capsules with a wide range of fluorescent dyes, including rhodamine 6G, pyronin B, fluorescein, acridine yellow, acridine orange, thiazine red, and previously reported rhodamine B. The fluorescence brightness, photostability, and photoconversion ability of the capsules are evaluated using confocal laser scanning microscopy. Viability, uptake, mobility, and photoconversion studies are conducted on RAW 264.7 and bone marrow-derived macrophages, serving as model cell lines. The production yield of the capsules is increased due to the use of polyvinyl alcohol gel, eliminating the need for conventional filtration steps. Capsules entrapping rhodamine B and rhodamine 6G meet all requirements for intracellular use in individual cell tracking. Mass spectrometry analysis reveals a sequence of deethylation steps that result in blue shifts in the dye spectra upon irradiation. Cellular studies on macrophages demonstrate robust uptake of the capsules. The capsules exhibit minimal cytotoxicity and have a negligible impact on cell motility. The successful photoconversion of RhB-containing capsules within cells highlights their potential as alternatives to photoconvertible proteins for individual cell labeling, with promising applications in personalized medicine.