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30 Aug 2002
German researchers improve the axial resolution of a confocal microscope by 34%.
Stefan Hell and Carlo Mar Blanca from Germany's Max Planck Institute of Biophysical Chemistry have improved the axial resolution of a confocal two-photon microscope to 330nm. This beats the previous resolution limit of 500 nm by 34%. (Optics Express 10 893)
The key component is a 1.45 numerical aperture oil-immersion lens made by Carl-Zeiss. "This gives the highest aperture angle, 72.8° available to immersion microscopy at present," report the authors in their paper. They combine the resolving power of the lens with a dark-ring (DR) binary aperture filter to further boost the resolution.
The researchers fire a beam of 760 nm light from a modelocked Ti:Sapphire laser through the specially designed DR filter. It divides the beam into two parts: a central maximum and a cylindrical outer portion. A careful design balances the amount of light in each portion.
As the beam passes through the filter, interference diverts part of the beam's energy away from the main maximum. According to Hell, this sharpens the maximum at the expense of spawning off higher-order lobes.
"When combining the DR filter and the NA=1.45 lens, we find experimentally that the main maximum is significantly sharpened up compared to that of a regular two-photon confocal microscope using a NA=1.4 lens," say the authors. "This is a new lower benchmark for single-lens imaging relying solely on fluorescence excitation."
Author
Jacqueline Hewett is news reporter on Optics.org and Opto & Laser Europe magazine.
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