Here, we report a β-galactosidase (β-Gal) responsive photochromic fluorescent probe NpG that was designed to pre-bind to human serum albumin (HSA) to form a probe/ protein hybrid NpG@HSA. The formation of NpG@HSA led to an increase in fluorescence emission (525 nm) corresponding to the binding of the fluorescent naphthalimide unit with HSA. In addition, this enabled visualization of the spiropyran fluorescence emission in aqueous media. Our probe/protein hybrid approach afforded a unique imaging platform with enhanced cell permeability and solubility that was capable of visualizing the cellular uptake of NpG@HSA before its activation by β-Gal. The β-Gal-mediated cleavage of the galactose unit within the NpG@HSA hybrid resulted in the formation of NpM@HSA and an increase in red fluorescence emission (620 nm). The resultant merocyanine unit was then able to undergo photoisomerization (merocyanine↔spiropyran) facilitating STORM (i.e. Stochastic Optical Reconstruction Microscopy) imaging with minimal phototoxicity and excellent photostability/reversibility. Using STORM, NpG@HSA was able to determine the subcellular distribution of β-Gal activity between cell lines with nanoscale precision. We believe this system represents a versatile imaging platform for the design of photochromic fluorescent probes suitable for illuminating the precise location of disease-specific biomarkers in various cellular processes.

中文翻译：

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用于生物重要生物标志物超分辨率成像的光致变色荧光探针策略

在这里，我们报告了一种 β-半乳糖苷酶 (β-Gal) 响应光致变色荧光探针 NpG，其设计用于与人血清白蛋白 (HSA) 预结合以形成探针/蛋白质杂交 NpG@HSA。NpG@HSA 的形成导致荧光发射（525 nm）增加，这与荧光萘酰亚胺单元与 HSA 的结合相对应。此外，这使得水介质中螺吡喃荧光发射的可视化成为可能。我们的探针/蛋白质杂交方法提供了一个独特的成像平台，具有增强的细胞渗透性和溶解性，能够在 β-Gal 激活之前可视化 NpG@HSA 的细胞摄取。NpG@HSA 杂交体中 β-Gal 介导的半乳糖单元裂解导致 NpM@HSA 的形成和红色荧光发射（620 nm）的增加。然后，所得部花青单元能够进行光异构化（部花青↔螺吡喃），促进 STORM（即随机光学重建显微镜）成像，同时具有最小的光毒性和出色的光稳定性/可逆性。使用 STORM，NpG@HSA 能够以纳米级精度确定细胞系之间 β-Gal 活性的亚细胞分布。我们相信该系统代表了一个通用的成像平台，用于设计适合照亮各种细胞过程中疾病特异性生物标志物的精确位置的光致变色荧光探针。NpG@HSA 能够以纳米级精度确定细胞系之间 β-Gal 活性的亚细胞分布。我们相信该系统代表了一个通用的成像平台，用于设计适合照亮各种细胞过程中疾病特异性生物标志物的精确位置的光致变色荧光探针。NpG@HSA 能够以纳米级精度确定细胞系之间 β-Gal 活性的亚细胞分布。我们相信该系统代表了一个通用的成像平台，用于设计适合照亮各种细胞过程中疾病特异性生物标志物的精确位置的光致变色荧光探针。