Super-resolution microscopy
Super-resolution microscopy comprises techniques that distinguish details smaller than the conventional diffraction-limited resolution of light microscopy. Many widely used methods employ fluorescence: selected molecules are labelled, and their emission is controlled or analysed to recover spatial information that would overlap in an ordinary image. These methods exploit additional information and different imaging conditions; they do not abolish the wave properties of light.[1]
Approaches
Structured illumination microscopy illuminates a specimen with a pattern. Images collected with different pattern positions and orientations allow fine structural information to be reconstructed computationally. Stimulated emission depletion, or STED microscopy, uses a second beam to suppress fluorescence around the centre of an excited region, leaving a smaller region contributing to the detected signal.[1]
Single-molecule localisation methods, including PALM and STORM, separate the emissions of nearby labels in time. The position of each isolated emitter can be estimated more precisely than the width of its blurred image. Repeated measurements of different subsets of molecules are combined into a map of their locations.[2]
Resolution and use
Locating one emitter precisely is not the same as resolving every feature in a densely labelled specimen. Image quality also depends on how well labels sample the structure, the number of detected photons and the accuracy of the reconstruction. The duration of data collection matters when the structure itself moves.[2]
Super-resolution methods have allowed cell biologists to examine the organisation of structures and molecular assemblies below the scale accessible to conventional fluorescence microscopy. The appropriate method depends on the experimental question, specimen preparation, available labels and the balance required between spatial detail and time resolution.[1]
References
- ↑ 1.0 1.1 1.2 Lothar Schermelleh, Rainer Heintzmann and Heinrich Leonhardt (2010). A guide to super-resolution fluorescence microscopy. Journal of Cell Biology.
- ↑ 2.0 2.1 Bo Huang, Mark Bates and Xiaowei Zhuang (2009). Super-resolution fluorescence microscopy. Annual Review of Biochemistry.
Attribution
Adapted with reference to English Wikipedia, revision 1375916484 (contributors and history), under CC BY-SA 4.0. The prose has been rewritten and checked against the cited sources; changes have been made.