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Journal Article

4Pi-confocal imaging in fixed biological specimens.

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Giese,  Günter
Department of Biomedical Optics, Max Planck Institute for Medical Research, Max Planck Society;

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Hell,  S. W.
Optical Nanoscopy, Max Planck Institute for Medical Research, Max Planck Society;

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Citation

Schrader, M., Bahlmann, K., Giese, G., & Hell, S. W. (1998). 4Pi-confocal imaging in fixed biological specimens. Biophysical Journal, 75(4), 1659-1668. doi:10.1016/S0006-3495(98)77608-8.


Cite as: https://hdl.handle.net/11858/00-001M-0000-0019-A44D-2
Abstract
By combining the wavefronts produced by two high-aperture lenses, two-photon 4Pi-confocal microscopy allows three-dimensional imaging of transparent biological specimens with axial resolution in the 100–140-nm range. We reveal the imaging properties of a two-photon 4Pi-confocal microscope as applied to a fixed cell. We demonstrate that a fast, linear point deconvolution suffices to achieve axially superresolved 3D images in the cytoskeleton. Furthermore, we describe stringent algorithms for alignment and control of the two lenses. We also show how to compensate for the effects of a potential refractive index mismatch of the mounting medium with respect to the immersion system.