Adaptive Optics Microscopy

Oscar Azucena (UCSC)

Changes in the index of refraction due to tissue composition limit the
resolving power of biological microscopy. This effect is more
pronounced in deep tissue imaging where the light travels through many
layers of cellular structures including cytoplasm and the plasma
membrane. Many important biological processes occur in deep tissue
such as stem cell division, neurogenesis and the key developmental
events following fertilization. A method that can be used to improve
deep tissue imaging is Adaptive Optics (AO). AO is a technique used in
astronomy to measure and correct the aberration introduced by
turbulence in the optical path. AO has also been applied to vision
science to enhance our understanding of the human eye. Two popular
techniques for improving resolution in optical systems are confocal
and two photon microscopy. These techniques also suffer from the
aberration introduced by the tissue and could be improved upon by
using AO. Most adaptive optics microscope systems so far have not
directly measured the wavefront due to the complexity of adding a
wavefront sensor in an optical system and the lack of a natural
point-source reference such as the “guide-star” used in astronomy and
vision science. Instead, most AO scanning microscopy systems have
corrected the wavefront by optimizing a signal received at a
photo-detector by using a hill-climbing algorithm. While there is a
lot of important research being done in AO microscopy, many of the AO
systems are specific to each microscope and a universal method for
measuring the wavefront (or the results of the correction algorithm)
is not currently available. A method for measuring the wavefront
aberrations induced by a Drosophila embryo by using a Shack-Hartmann
wavefront sensor and light emitted from an imbedded florescent
microsphere will be presented. The Drosophila embryo is well suited to
analysis as it is approximately 200 μm in diameter, rich in cytoplasm
and amenable to experimental manipulation.:wq