RT2-01962 Adaptive Optics Two-Photon Microscope for Single Cell Imaging of Stem Cells Embedded in Deep Tissue

Funding Type: 
Tools and Technologies II
Grant Number: 
RT2-01962
Investigator: 
ICOC Funds Committed: 
$0
oldStatus: 
Closed
Public Abstract: 
To speed the delivery of stem cell therapies from research in the laboratory to practice in the clinic, our collaborative research team composed of biologists, physicists, astronomers and engineers proposes to develop a new two-photon microscope with Adaptive Optics (AO) that can see more clearly when imaging deeply into the brain. Adaptive optics are used by astronomers to provide clearer images of stars. In astronomy, lasers are used to create reference beacons which can be used to adjust the optics in the telescope to correct for image distortions caused by changes in the atmosphere, such as winds and dynamic temperature changes that cause the stars to “twinkle.” Such distortions can then be corrected by using a dynamically deformable mirror, similar to the curved mirrors in fun houses, which make you look short or tall. In principle, reference beacons and deformable mirrors can also be used to improve image depth and resolution of stem cell samples when looking through thick tissue. Here the reference beacons control the mirrors that remove warping of the image caused by looking through the materials inside the cell. Our preliminary results indicate small fluorescent beads can be implanted in tissue to serve as beacons to acquire the necessary measurements to correct for most distortions. The new microscope that will be developed uses two photons to excite fluorescence, which enables it to image more deeply into tissue. The new two-photon AO microscope builds upon our prior success using adaptive optical techniques for a single-photon AO microscope which has been shown to be capable of improving the image resolution by ten times the resolution of conventional microscopes. Many promising stem cell-based therapies will require injecting in-vitro grown stem cells into a specific afflicted tissue. However, previous attempts at reviving or repairing damaged tissues by injecting stem cells have had limited success. The current lack of understanding of the cellular basis of this failure has severely impeded advancement of such therapies. A major technical barrier has been the limitation for visualizing individual living stem cells in deep tissue, thus preventing investigators from answering many critical questions. For example, do such injected cells continue to undergo normal stages of self renewal and differentiation? Once the stem cells migrate to particular niches, what is the fate of stem cells that subsequently differentiate? We will use this new microscope for tracking transplanted stem cell derivatives within a living organism to study how stem cells can help repair brain damage after a stroke. By using adaptive optics we will be able to increase the imaging depth and resolution to near diffraction limited imaging.
Statement of Benefit to California: 
Successful development of an adaptive optics two-photon microscope will have an immediate benefit to stem cell research, particularly that research funded by the California Institute for Regenerative Medicine (CIRM). Stem cells usually reside deep within dense cell layers and thus have proven extremely difficult for live fluorescent imaging. Live fluorescent imaging has become an essential tool for revealing the molecular and cell biology of dynamic biological events. With respect to stem cell biology, live cellular imaging will be essential to address the translational bottleneck for the development of sensitive imaging and molecular techniques for tracking transplanted stem cell derivatives in vivo. Many promising stem cell-based therapies will require injecting in vitro grown stem cells into a specific afflicted tissue. However, previous attempts at reviving or repairing damaged tissues by injecting stem cells have had limited success. The current lack of understanding of the cellular basis of this failure has severely impeded advancement of such therapies. A major technical barrier has been the limitation for visualizing individual living stem cells in deep tissue, thus preventing investigators from answering many critical questions. The adaptive optics two-photon microscope we are developing should enable us to perform live fluorescent imaging at depths more than double the current limit of 250 microns. We hope that within the decade AO two-photon microscopes will be a standard tool for stem cell biologists. We will use this new microscope for tracking transplanted stem cell derivatives within a living organism to study how stem cells can help repair brain damage after a stroke. In addition, it will have general applicability for treatment of many other neurological diseases. Although two-photon microscopy is best applied to tracking transplanted stem cells in small animal models, the results will be important for understanding disease in humans. We believe this project will also have direct benefit to the California economy as the manufacturing, distribution and selling of the microscope would take place in California. We have already had interest from local optics companies in developing the AO microscope for market once a suitable prototype is developed.

© 2013 California Institute for Regenerative Medicine