Robinson Research Institute and Centre for Nanoscale BioPhotonics, Adelaide Medical School, University of Adelaide , Adelaide, South Australia 5005, Australia.
Institute of Molecular Medicine, Sechenov First Moscow State University , Moscow 119991, Russia.
ACS Appl Mater Interfaces. 2017 Nov 15;9(45):39197-39208. doi: 10.1021/acsami.7b12665. Epub 2017 Nov 2.
At the forefront of developing fluorescent probes for biological imaging applications are enhancements aimed at increasing their brightness, contrast, and photostability, especially toward demanding applications of single-molecule detection. In comparison with existing probes, nanorubies exhibit unlimited photostability and a long emission lifetime (∼4 ms), which enable continuous imaging at single-particle sensitivity in highly scattering and fluorescent biological specimens. However, their wide application as fluorescence probes has so far been hindered by the absence of facile methods for scaled-up high-volume production and molecularly specific targeting. The present work encompasses the large-scale production of colloidally stable nanoruby particles, the demonstration of their biofunctionality and negligible cytotoxicity, as well as the validation of its use for targeted biomolecular imaging. In addition, optical characteristics of nanorubies are found to be comparable or superior to those of state-of-the-art quantum dots. Protocols of reproducible and robust coupling of functional proteins to the nanoruby surface are also presented. As an example, NeutrAvidin-coupled nanoruby show excellent affinity and specificity to μ-opioid receptors in fixed and live cells, allowing wide-field imaging of G-protein coupled receptors with single-particle sensitivity.
在开发用于生物成像应用的荧光探针方面，人们致力于提高其亮度、对比度和光稳定性，特别是在单分子检测等苛刻的应用中。与现有探针相比，纳米棒具有无限的光稳定性和较长的发射寿命（约 4 毫秒），这使其能够在高度散射和荧光生物样本中以单粒子灵敏度进行连续成像。然而，由于缺乏大规模生产和分子特异性靶向的简便方法，纳米棒在荧光探针中的广泛应用受到了限制。本研究包括胶体稳定的纳米棒颗粒的大规模生产，证明了其生物功能和可忽略的细胞毒性，并验证了其用于靶向生物分子成像的用途。此外，纳米棒的光学特性被发现可与最先进的量子点相媲美。还提出了可重现且稳定的将功能蛋白偶联到纳米棒表面的方案。例如，与 NeutrAvidin 偶联的纳米棒对固定和活细胞中的 μ 阿片受体表现出优异的亲和力和特异性，允许用单粒子灵敏度对 G 蛋白偶联受体进行宽场成像。
