Department of Applied Physics and SciLifeLab, KTH Royal Institute of Technology, 114 28, Stockholm, Sweden.
Chemphyschem. 2023 Feb 1;24(3):e202200698. doi: 10.1002/cphc.202200698. Epub 2022 Nov 9.
Stimulated emission depletion (STED) nanoscopy is a widely used nanoscopy technique. Two-colour STED imaging in fixed and living cells is standardised today utilising both fluorescent dyes and fluorescent proteins. Solutions to image additional colours have been demonstrated using spectral unmixing, photobleaching steps, or long-Stokes-shift dyes. However, these approaches often compromise speed, spatial resolution, and image quality, and increase complexity. Here, we present multicolour STED nanoscopy with far red-shifted semiconductor CdTe quantum dots (QDs). STED imaging of the QDs is optimized to minimize blinking effects and maximize the number of detected photons. The far-red and compact emission spectra of the investigated QDs free spectral space for the simultaneous use of fluorescent dyes, enabling straightforward three-colour STED imaging with a single depletion beam. We use our method to study the internalization of QDs in cells, opening up the way for future super-resolution studies of particle uptake and internalization.
受激发射耗散（STED）纳米显微镜是一种广泛应用的纳米显微镜技术。今天，利用荧光染料和荧光蛋白，已经实现了双色 STED 成像在固定和活细胞中的标准化。已经证明了使用光谱解混、光漂白步骤或长斯托克斯位移染料来实现额外颜色成像的解决方案。然而，这些方法通常会降低速度、空间分辨率和图像质量，并增加复杂性。在这里，我们展示了使用远红色移位的半导体 CdTe 量子点（QD）的多色 STED 纳米显微镜。QD 的 STED 成像经过优化，以最小化闪烁效应并最大化检测到的光子数量。所研究的 QD 的远红和紧凑的发射光谱为荧光染料的同时使用提供了自由光谱空间，从而可以使用单个耗尽光束进行简单的三色 STED 成像。我们使用我们的方法研究了 QD 在细胞中的内化，为未来对颗粒摄取和内化的超分辨率研究开辟了道路。
