Non-destructive fluorophore diffusion across cell membranes to provide an unbiased fluorescence intensity readout is critical for quantitative imaging applications in live cells and tissues. Commercially available small-molecule fluorophores have been engineered for biological compatibility, imparting high water solubility by modifying rhodamine and cyanine dye scaffolds with multiple sulfonate groups. The resulting net negative charge, however, often renders these fluorophores cell-membrane-impermeant. Here we report the design and development of our biologically compatible, water-soluble and cell-membrane-permeable fluorophores, termed OregonFluor (ORFluor). By adapting previously established ratiometric imaging methodology using bio-affinity agents, it is now possible to use small-molecule ORFluor-labelled therapeutic inhibitors to quantitatively visualize their intracellular distribution and protein target-specific binding, providing a chemical toolkit for quantifying drug target availability in live cells and tissues.

跨细胞膜的非破坏性荧光团扩散以提供无偏差的荧光强度读数对于活细胞和组织的定量成像应用至关重要。市售小分子荧光团经过精心设计，具有生物相容性，通过用多个磺酸基团修饰罗丹明和花青染料支架来赋予高水溶性。然而，由此产生的净负电荷通常使这些荧光团不能渗透细胞膜。在这里，我们报告了生物相容性、水溶性和细胞膜可渗透荧光团的设计和开发，称为 OregonFluor (ORFluor)。通过采用先前建立的使用生物亲和剂的比例成像方法，现在可以使用小分子 ORFluor 标记的治疗抑制剂来定量可视化其细胞内分布和蛋白质靶标特异性结合，从而提供化学工具包来量化药物靶标的可用性活细胞和组织。