DR-NANO · Depth-Resolved Optical Nanoscopy
FP7 — People (Marie Curie Actions)
- Duration
- 2011-09-01 → 2015-08-31
- EU contribution
- €100,000
- Participants
- 1
- Scheme
- MC-CIG
Lines connect the coordinator with its partners.
Results in brief
Depth-Resolved Optical Nanoscopy
The overall objective of the project is to develop a fluorescence microscope for sub-diffraction-limit visualization in thick samples. The microscope is based on the illumination of a confined layer in the specimen and 4pi detection of the emitted fluorescence to localize fluorescent probes in three dimensions from specific layers in the sample. The anticipated use of the microscope is in the characterization of the nanoenvironment of soft bio-materials and in biological imaging. Since the beginning of the project, the illumination and detection paths of the microscope have been developed and optimized and depth-resolved, two-dimensional imaging and tracking of fluorescent probes in thick bio-gels and F-actin solutions have been demonstrated using a unique multiple particle tracking analysis framework developed in the project. Furthermore, the use of photoactivatable fluorescent molecules with the proposed microscope and the molecular localization algorithm developed in the project has been tested for nano-imaging in thick samples. Finally, the performance of the 4pi detection path of the microscope for axial localization has been explored experimentally and has shown a 13-nm axial localization precision, presenting a promising path towards sub-diffraction-limit imaging and tracking in thick samples in three dimensions. The developed microscope presents a new investigational tool for the analysis of the nanoenvironment of complex material and it is expected that the techniques and knowledge gained throughout the project will promote many other studies of sub-diffraction-limit visualization deep in biological and physical systems as well as will contribute to European excellence and European competitiveness in this important field of optical nano-imaging.
Data: CORDIS, © European Union
Project objective
A fluorescence microscope for the noninvasive imaging of the structure and dynamics in thick 3D systems with 3D sub-diffraction-limited resolution and at extended penetration depths would have a broad spectrum of applications in both the physical and biological sciences. Conventional fluorescence microscopy techniques, such as confocal and two-photon microscopy, lack the spatial resolution required for measurements at the nanoscale, whereas state-of-the-art 3D super-resolution optical microscopy methods offer limited penetration depth (<5 microns). In the proposed project, I aim to develop a new microscope that combines molecular photoswitches, fluorescence self-interference and light-sheet microscopy to optically section a thin layer deep in thick semitransparent samples with 3D sub-diffraction-limited resolution. I will then test it for potential physical and biological applications of single molecule detection. The first specific aim is to develop a depth-resolved self-interference photoswitching nanoscope employing novel high-throughput and sub-diffraction-limit fluorescence interferometry. The second specific aim is to map with 3D nanometer resolution the motion of photoswitchable fluorescent probes deep in soft materials. This study will open up new possibilities for precise measurements of the heterogeneity and mechanical properties of the nanoenvironments of soft matter at extended depths and may ultimately assist in developing superior biomaterials and nanomedical therapeutics. The third specific aim is to in situ image with 3D nanometer resolution the muscle thick filament system in fixed Caenorhabditis elegans nematodes at both the ventral and dorsal ends (separated by ~80 microns). This demonstration, if successful, will provide unique 3D super-resolution in situ image data and may assist in developing imaging protocols for in situ nanoscopy of structural changes of the C. elegans body-wall muscle system due to sarcomeric muscle diseases.
Original text from CORDIS.
Participants
- BEN-GURION UNIVERSITY OF THE NEGEV · Beer ShevaCoordinatorIsrael
Links
Data: CORDIS, © European Union