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The image resolution and contrast of microscopes are often detrimentally affected by aberrations that are introduced when focusing deep into specimens. These aberrations arise from spatial differences in optical properties of the specimen or refractive index mismatches. This is particularly problematic in multiphoton microscopy, where short pulsed lasers are used to generate contrast through non-linear optical effects, such as two-photon fluorescence or third harmonic generation. The non-linear nature of the signal generation process means that the signal level is strongly affected by changes in the focal spot intensity. We have applied the techniques of adaptive optics to measure and correct the aberrations, restoring image quality. In particular, this has been demonstrated in harmonic generation microscopy of developing mouse embryos. Similar aberration problems affect the resolution and efficiency of three-dimensional optical fabrication systems, such those used for the manufacture of photonic crystals or optical waveguides. These systems are based around microscope optics and use short pulsed laser illumination to induce localized multiphoton effects in a fabrication substrate. In this case, significant aberrations are introduced when focusing deep into the substrate. We report on the development of adaptive optics systems for these applications and discuss the specific challenges for wave front sensing and correction that are presented by these systems. © 2010 Copyright SPIE - The International Society for Optical Engineering.

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