🏆 Foundational Paper

Super-resolution fluorescence imaging with single molecules.

Sahl Steffen J, Moerner W E

📰 Current opinion in structural biology 📅 2013 📊 143 citations
DOI PubMed

Abstract

The ability to detect, image and localize single molecules optically with high spatial precision by their fluorescence enables an emergent class of super-resolution microscopy methods which have overcome the longstanding diffraction barrier for far-field light-focusing optics. Achieving spatial resolutions of 20-40nm or better in both fixed and living cells, these methods are currently being established as powerful tools for minimally-invasive spatiotemporal analysis of structural details in cellular processes which benefit from enhanced resolution. Briefly covering the basic principles, this short review then summarizes key recent developments and application examples of two-dimensional and three-dimensional (3D) multi-color techniques and faster time-lapse schemes. The prospects for quantitative imaging - in terms of improved ability to correct for dipole-emission-induced systematic localization errors and to provide accurate counts of molecular copy numbers within nanoscale cellular domains - are discussed.

🔬 Techniques

DNA Points Accumulation for Imaging in Nanoscale Topography Fluorescence Microscopy Immunofluorescence Microscopy Live Cell Imaging Optical Sectioning Photoactivated Localization Microscopy Reversible Saturable Optical Fluorescence Transitions Microscopy Single-Molecule Imaging Stimulated Emission Depletion Microscopy Stochastic Optical Reconstruction Microscopy Super-Resolution Microscopy Total Internal Reflection Fluorescence Microscopy Widefield Fluorescence Microscopy

🧬 Organisms

Escherichia coli Xenopus laevis

✨ Fluorophores

Cy3 Cy5 Dendra2 EYFP mEos2 YFP

🧪 Sample Preparation

Fixation Immunofluorescence Immunostaining Live Imaging

🔬 Cell Lines

Chinese Hamster Ovary Human Osteosarcoma U2OS Rat Pheochromocytoma PC-12

📷 Detectors

Photometrics evolve

🏛️ Research Organizations (ROR)

Affiliated research institutions:

🏛 Stanford University 🏛 The Classical Association

📊 Figures

Figure 1

Figure 1

Principles of super-resolution single-molecule active control microscopy. (a) A hypothetical arrangement of fluorescent molecules, i.e. a labeled u201csuper-structureu201d (here: outline of u201cLa Pa...

Figure 2

Figure 2

Examples of single-molecule-based super-resolution imaging in biological applications. (a) The nucleoid-associated protein HU imaged at different stages in the cell cycle of asymmetrically dividing C....

Figure 3

Figure 3

Three-dimensional (3D) super-resolution fluorescence imaging. (a) To extract the location of individual single-molecule emitters with high precision in all three spatial dimensions, the symmetry of th...

Figure images are served from the NIH/NLM PubMed Central Open Access Subset or Europe PMC; copyright remains with the publishers and authors.

🏛️ Imaging Facility

🏛️ Stanford University

💬 Discussion

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