What if deep tissue images were continuously sharp, from the coverslip down to hundreds of microns into the sample?
Our Adaptive Optics system based on a deformable mirror does exactly that: by correcting the refractive index mismatch between immersion and embedding media, plus sample-induced aberrations, your images are crisp and bright everywhere. Even for very large focusing depths, with any objective lens, and any embedding medium.
From confocal microscopy over multiphoton imaging to 2D- and 3D-STED, all techniques suﬀer from aberrations that inevitably occur when focusing into a sample whose refractive index is diﬀerent from the immersion medium, or when focusing into a sample with internal inhomogeneities.
Using a deformable mirror that shapes the beam so as to pre-compensate for aberrations, all rays are put back in the right place for a perfectly crisp focus.
Spherical aberrations arising from index mismatch between sample and immersion are corrected for, as well as higher-order, sample-induced aberrations.
Without Adaptive Optics, excitation laser power is typically increased with focus depth in order to compensate loss of signal due to aberrations. Adaptive Optics preserves resolution and brightness deep inside thick samples and enables imaging at low light levels (sample courtesy of Amelie Cabirol and Albrecht Haase, University of Trento).
With Adaptive Optics, we enhance the performance of our microscopes by manipulating the wavefronts of the STED, excitation and detection beams using a deformable mirror.
Undistorted wavefronts entering the sample from the objective lens are normally spherical, but variations of the refractive index in the sample can distort them, leading to an imperfect focus. The prime reason, which is almost always present to some extent, is a refractive index mismatch between sample embedding and the immersion medium, although local variations in the sample can lead to aberrations, too.
Using a deformable mirror allows us to effectively cancel aberrations. Deformable mirrors are adaptive elements with a reflective surface whose shape can be controlled. By applying the correct mirror shape, which is a negative of the distortions introduced by the sample, the focus is brought back to perfect shape, increasing signal and resolution even deep inside tissue.