研究背景：仍然存在一些挑战，如在不同肿瘤中开发用于肿瘤边缘鉴定的通用肿瘤特异性探针，且具有易操作和快速成像模式。因此，本研究构建了一个具有酸激活荧光和大斯托克斯移动的快速“开-关”近红外（NIR）荧光探针NBD，用于喷雾介导的近即时和精确的肿瘤边缘鉴定。方法：通过在β-咔啉的C-6和C-3位置引入二苯胺基团和苯并[丁]吲哚基团以设计和合成NBD。通过吸收光谱、荧光光谱对NBD的光学性质进行表征。随后，我们通过1H NMR和密度泛函理论（DFT）计算研究了其pH依赖机制。进一步深入研究了NBD在裸鼠模型（皮下、正位和转移瘤）和来自三种临床代表性肿瘤（肝癌、结肠癌和肺癌）患者的临床组织中的成像性能。结果：发现NBD具有近红外荧光（742 nm）、大斯托克斯移动（160 nm）和双光子吸收（1040 nm）。荧光量子产率（ФF）在pH从7.4降至4.0时增加了5.5倍，表现出pH依赖性。此外，NBD不仅能选择性地照亮四株癌细胞系，还能勾勒出异种移植瘤和正位微小肿瘤，用于指导外科肿瘤切除，并追踪转移组织。特别是，在简单的局部喷洒后（三分钟后），NBD能够快速而准确地区分包括肝癌、结肠癌和肺癌在内的三种临床癌症标本的边界范围，具有较高的肿瘤与正常组织信号比（6.48〜9.80）。结论：因此，所提出的荧光探针NBD可能作为一种通用的近红外荧光喷雾，用于近即时可视化肿瘤边缘并帮助外科医生进行临床癌症手术切除。©作者。

Rationale: Challenges such as developing a universal tumor-specific probe for tumor margin identification in diverse tumors with an easy-operative and fast-imaging pattern still exist. Hence, in the present study, a rapidly "off-on" near-infrared (NIR) fluorescent probe NBD with pH-activatable fluorescence and a large Stokes shift was constructed for spray mediated near-instant and precise clinical tumor margins identification. Methods: NBD was designed and synthesized by introducing both diphenyl amino group and benzo[e]indolium to β-carboline at C-6 and C-3 positions respectively. The optical properties of NBD was characterized by absorption spectra, fluorescence spectra. Subsequently, we investigated its pH-dependent mechanism by 1H NMR and density functional theory (DFT) calculation. NBD was further under deeper investigation into its imaging performance in nude mice models (subcutaneous, orthotopic, metastatic tumor), and clinical tissues from patients with three clinically representative tumors (liver cancer, colon cancer, and lung cancer). Results: It was found that NBD had NIR fluorescence (742 nm), a large Stokes shift (160 nm), and two-photon absorbance (1040 nm). Fluorescence quantum yield (ФF) increased by 5.5-fold when pH decreased from 7.4 to 4.0, to show pH-dependent property. Furthermore, NBD could not only selectively light up all four cancer cell lines, but also delineate xenograft tumor and orthotopic microtumor to guide surgical tumor resection, and track metastatic tissues. Particularly, after simple topical spray (three minutes later), NBD could rapidly and precisely distinguish the boundary ranges of three kinds of clinical cancer specimens including liver, colon, and lung cancers, with high tumor-to-normal tissue signal ratios (6.48~9.80). Conclusions: Therefore, the proposed fluorescent probe NBD may serve as a versatile NIR fluorogenic spray for the near-instant visualization of tumor margins and assisting surgeons in surgerical resection of clinical cancers.© The author(s).