MRC-Laboratory for Molecular Cell Biology, University College London, London, UK.
The Francis Crick Institute, London, UK.
Traffic. 2020 May;21(5):375-385. doi: 10.1111/tra.12728. Epub 2020 Mar 30.
Localization-based super-resolution microscopy relies on the detection of individual molecules cycling between fluorescent and non-fluorescent states. These transitions are commonly regulated by high-intensity illumination, imposing constrains to imaging hardware and producing sample photodamage. Here, we propose single-molecule self-quenching as a mechanism to generate spontaneous photoswitching. To demonstrate this principle, we developed a new class of DNA-based open-source super-resolution probes named super-beacons, with photoswitching kinetics that can be tuned structurally, thermally and chemically. The potential of these probes for live-cell compatible super-resolution microscopy without high-illumination or toxic imaging buffers is revealed by imaging interferon inducible transmembrane proteins (IFITMs) at sub-100 nm resolutions.
基于定位的超分辨率显微镜依赖于检测在荧光和非荧光状态之间循环的单个分子。这些转变通常受到高强度照明的调节，对成像硬件提出了限制，并产生了样品光损伤。在这里，我们提出单分子自猝灭作为产生自发光开关的机制。为了证明这一原理，我们开发了一类新的基于 DNA 的开源超分辨率探针，命名为超级灯塔，其光开关动力学可以通过结构、热和化学进行调节。通过以亚 100nm 的分辨率对干扰素诱导的跨膜蛋白（IFITMs）进行成像，揭示了这些探针在没有高强度照明或有毒成像缓冲液的情况下用于活细胞兼容的超分辨率显微镜的潜力。
