PDF(Pubmed)
Abstract：
Molecular probes with the ability to differentiate between subcellular variations in acidity levels remain important for the investigation of dynamic cellular processes and functions. In this context, a series of cyclic peptide and PEG bio-conjugated dual near-infrared emissive BF2-azadipyrromethene fluorophores with maxima emissions at 720 nm (at pH > 6) and 790 nm (at pH < 5) have been developed and their aqueous solution photophysical properties determined. Their inter-converting emissions and fluorescence lifetime characteristics were exploited to track their spatial and temporal progression from first contact with the plasma membrane to subcellular locales to their release within extracellular vesicles. A pH-dependent reversible phenolate/phenol interconversion on the fluorophore controlled the dynamic changes in dual emission responses and corresponding lifetime changes. Live-cell confocal microscopy experiments in the metastatic breast cancer cell line MDA-MB-231 confirmed the usability of the dual emissive properties for imaging over prolonged periods. All three derivatives performed as probes capable of real-time continuous imaging of fundamental cellular processes such as plasma membrane interaction, tracking endocytosis, lysosomal/large acidic vesicle accumulation, and efflux within extracellular vesicles without perturbing cellular function. Furthermore, fluorescence lifetime imaging microscopy provided valuable insights regarding fluorophore progression through intracellular microenvironments over time. Overall, the unique photophysical properties of these fluorophores show excellent potential for their use as information-rich probes.
摘要：
能够区分酸度水平的亚细胞变化的分子探针对于研究动态细胞过程和功能仍然很重要。在这种情况下，已经开发了一系列环状肽和PEG生物缀合的双近红外发射BF2-氮杂二吡咯亚甲基荧光团,其在720nm(在pH>6下)和790nm(在pH<5下)具有最大发射,并且确定了它们的水溶液光物理性质。利用它们的相互转换发射和荧光寿命特征来跟踪它们从与质膜的首次接触到亚细胞区域到它们在细胞外囊泡内的释放的空间和时间进程。荧光团上的pH依赖性可逆的酚盐/酚相互转化控制了双重发射响应的动态变化和相应的寿命变化。转移性乳腺癌细胞系MDA-MB-231中的活细胞共聚焦显微镜实验证实了双重发射特性在长时间成像中的可用性。所有这三种衍生物都是作为探针进行的，能够实时连续成像基本细胞过程，如质膜相互作用，追踪内吞作用，溶酶体/大酸性囊泡积累，并在细胞外囊泡内流出而不干扰细胞功能。此外，荧光寿命成像显微镜提供了有关荧光团通过细胞内微环境随着时间的推移进展的有价值的见解。总的来说,这些荧光团独特的光物理性质显示出作为信息丰富的探针的巨大潜力。
