Design of an AO microscope for biological imaging
Creators: name: University Of California Office Of The President
Publisher: name: University of California Office of the President
publisherIdentifier: https://ror.org/00dmfq477
publisherIdentifierScheme: ROR
Year: 2024

description: While a new generation of microscopes and fluorescent probes has revolutionized live imaging of biological specimens, this technology cannot be applied to deep-tissue imaging of living cells. With only a few exceptions, at depths greater than 30 microns, the quality of live imaging is poor. For example although the early cellular and developmental events of the Drosophila embryo are intensively investigated by thousands of researchers around the world, we still do not have live images of the key events immediately following fertilization. Much of the degradation in image quality is the result of local differences in the refractive index, both within the sample and between the sample and the lens. The proposed project is for the design of a wide-field and a confocal adaptive optic (AO) microscope for deep-tissue imaging of living cells using reference beacons, wavefront sensing and correction. Based on related AO results in vision science, this approach could potentially increase the Strehl ratio of the image from less than 2% to over 70%. The proposed two-year activity will transform knowledge and understanding within disciplines ranging from cell biology and genetics to microbiology and developmental biology by increasing the depth of high-resolution imaging into tissue where many fundamental cellular processes occur. Examples include the fusion of male and female genomes after fertilization; neuron growth, organization and synapse formation; formation of the germline, which is responsible for sperm and egg production; and fate mapping of mammalian embryonic stem cells. The design team, with members from UCSC, UCSF and LLNL, is extremely well-qualified to conduct the project and have already been working together under seed funding from the Center for Adaptive Optics to obtain preliminary wavefront aberration data required for the design. Members of the team implemented the first astronomical AO systems using artificial laser guide stars at the Lick and Keck observatories, transferred the resulting AO technology into the field of vision science, and are currently working on the next generation of high-stroke, high-order Micro-Electro-Mechanical (MEMS)-based AO systems in both of these fields. While other research programs are using adaptive optics in microscopy, there has been no work using reference beacons and wavefront sensing with AO to directly measure the physical quantity that is corrected by the deformable mirror, as we propose in our microscope design. This approach has been successfully demonstrated and adopted as the standard in most fields in which AO is used, but has never been implemented in biological imaging. The proposed design is based on low-risk modifications to the proven LLNL Adaptive Optics Scanning Laser Ophthalmoscope (AOSLO) and the Adaptive Optics Wide-Field Microscope (AOWFM), and includes team members who were instrumental in designing and building the highest resolution MEMS-based adaptive optic microscopes to date. Available resources include UCSC's state-of-the-art Laboratory for Adaptive Optics (LAO), the first and only comprehensive laboratory in the United States dedicated to adaptive optics, and a leader in the development of MEMS deformable mirrors for adaptive optics. The LAO also provides an opportunity for both undergraduate and graduate students to become involved in the development of multidisciplinary, adaptive optics based instruments.