Spatially modulated illumination microscopy using one objective lens

Hans Mathée; David Baddeley; C. Wotzlaw; Christoph G. Cremer; Udo J. Birk

doi:10.1117/1.2768085

1 August 2007 Spatially modulated illumination microscopy using one objective lens

Hans Mathée, David Baddeley, C. Wotzlaw, Christoph G. Cremer, Udo J. Birk

Author Affiliations +

Optical Engineering, Vol. 46, Issue 8, 083603 (August 2007). https://doi.org/10.1117/1.2768085

Abstract

The spatially modulated illumination (SMI) microscope is a wide-field fluorescence microscope featuring axially structured illumination, through which access to information about subresolution object structures is obtained. We present a simplified setup where the interference pattern is generated by reflecting the laser beam with a mirror. We characterize our setup by presenting measurements on fluorescent microspheres with diameters ranging from 44 to 200 nm. The results agree well with the sizes provided by the manufacturer. Furthermore, the spheres are analyzed with 458-, 514-, 488-, and 568-nm excitation wavelengths, giving good agreement of the sizes determined at the respective wavelengths. A measurement of the same objects using different excitation wavelengths leads to a size difference of a few nanometers only. The potential of SMI microscopy for the fast analysis of many fluorescent objects is also addressed. In addition, the applicability to biological specimens is shown on fluorescently labeled, specific chromatin domains. The results obtained using the presented mirror geometry agree well with data obtained using a standard SMI microscope setup.

©(2007) Society of Photo-Optical Instrumentation Engineers (SPIE)

Citation Download Citation

Hans Mathée, David Baddeley, C. Wotzlaw, Christoph G. Cremer, and Udo J. Birk "Spatially modulated illumination microscopy using one objective lens," Optical Engineering 46(8), 083603 (1 August 2007). https://doi.org/10.1117/1.2768085

Published: 1 August 2007

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