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Multiphoton (MP) microscopy is a well-established method for the non-invasive imaging of biological tissues. However, its optical sectioning capabilities are reduced due to specimen-induced aberrations. Both the manipulation of spherical aberration (SA) and the use of axicons have been reported to be useful techniques to bypass this limitation. We propose the combination of SA patterns and variable axicons to further improve the quality of MP microscopy images. This approach provides enhanced images at different depth locations whose quality is better than those corresponding to the use of SA or axicons separately. Thus, the procedure proposed herein facilitates the visualization of details and increases the depth observable at high resolution.

Improving Multiphoton Microscopy by Combining Spherical Aberration Patterns and Variable Axicons