Compact 4pi beam scanning microscope for imaging cells with a sevenfold increase in axial resolution

Hilmar F. Gugel; Reiner Rygiel; Rafael Storz

doi:10.1117/12.618525

30 March 2005 Compact 4pi beam scanning microscope for imaging cells with a sevenfold increase in axial resolution

Hilmar F. Gugel, Reiner Rygiel, Rafael Storz

Proceedings Volume 5700, Multiphoton Microscopy in the Biomedical Sciences V; (2005) https://doi.org/10.1117/12.618525
Event: SPIE BiOS, 2005, San Jose, CA, United States

Abstract

Confocal microscopy is the method of choice in biological 3D-imaging, however, the axial resolution is limited to ~500 nm. During the last decade it has been successfully demonstrated that the axial resolution can be substantially improved with 4Pi microscopy. We report a 4Pi microscope realized as a fast beam scanning system consisting of a 4Pi-module linked to a state-of-the-art confocal microscope. As a result, the advantages of the confocal system such as scanning, sensitive multicolor detection and imaging speed are combined with the superior resolution of 4Pi microscopy. This novel microscope is eminently suited for biological applications. It is designed both for single-photon and for two-photon picosecond excitation, and also enables joint coherent illumination and detection, i.e. 4Pi type C. The superior PSF and OTF of the system enable 80 nm axial resolution in cells mounted in aqueous media. We present the optical design of the system and demonstrate an up to 7-fold improved optical sectioning in live cells.

Citation Download Citation

Hilmar F. Gugel, Reiner Rygiel, and Rafael Storz "Compact 4pi beam scanning microscope for imaging cells with a sevenfold increase in axial resolution", Proc. SPIE 5700, Multiphoton Microscopy in the Biomedical Sciences V, (30 March 2005); https://doi.org/10.1117/12.618525

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