Resolution enhancement in a light-sheet-based microscope (SPIM)

CJ Engelbrecht, EH Stelzer - Optics letters, 2006 - opg.optica.org
CJ Engelbrecht, EH Stelzer
Optics letters, 2006opg.optica.org
Light-sheet-based microscopy [single-plane illumination microscope (SPIM)] performs very
well at low numerical apertures. It complements conventional (FM), confocal (CFM), and two-
photon fluorescence microscopy (2hν-FM) currently used in modern life sciences. Lateral
and axial SPIM point spread function (PSF) extents are measured by using fluorescent
beads to determine the 3D resolution. The results are compared with values derived from an
analytical theory and numerical simulations. The discrepancies are found to be less than …
Light-sheet-based microscopy [single-plane illumination microscope (SPIM)] performs very well at low numerical apertures. It complements conventional (FM), confocal (CFM), and two-photon fluorescence microscopy (2hν-FM) currently used in modern life sciences. Lateral and axial SPIM point spread function (PSF) extents are measured by using fluorescent beads to determine the 3D resolution. The results are compared with values derived from an analytical theory and numerical simulations. The discrepancies are found to be less than 5%. The axial extent of a SPIM-PSF (10×/0.3 W) is approximately 5.7 μm. This value is almost a factor of 2 smaller than in CFM, more than 2.5 times smaller than in FM, and more than three times smaller than in 2hν-FM. SPIM outperforms 2hν-FM and FM, while CFM has a better axial resolution at NAs above 0.8.
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