Breaking the diffraction barrier in fluorescence microscopy by optical shelving.

Journal article

We report the breaking of the diffraction resolution barrier in far-field fluorescence microscopy by transiently shelving the fluorophore in a metastable dark state. Using a relatively modest light intensity of several kW/cm(2) in a focal distribution featuring a local zero, we confine the fluorescence emission to a spot whose diameter is a fraction of the wavelength of light. Nanoscale far-field optical resolution down to 50 nm is demonstrated by imaging microtubules in a mammalian cell and proteins on the plasma membrane of a neuron. The presence of dark states in virtually any fluorescent molecule opens up a new venue for far-field microscopy with resolution that is no longer limited by diffraction.

Authors: Bretschneider, S; Eggeling, C; Hell, S

• Journal: Physical Review Letters
• Volume: 98
• Issue: 21
• Pages: 218103
• Publication date: 2007-05-01
• DOI: 10.1103/physrevlett.98.218103
• EISSN: 1079-7114
• ISSN: 0031-9007
• Language: English
• Keywords: Humans; Microscopy, Fluorescence; Optics and Photonics; Cell Line; Synaptosomal-Associated Protein 25; Fluorescent Dyes