Hyperspectral stimulated emission depletion microscopy and methods of use thereof
| DWPI Title: Method for multi-color and sub-diffraction imaging of three-dimensional space, involves providing sample that includes three-dimensional space having features of interest, which are smaller than optical diffraction limit |
| Abstract: A hyperspectral stimulated emission depletion (“STED”) microscope system for high-resolution imaging of samples labeled with multiple fluorophores (e.g., two to ten fluorophores). The hyperspectral STED microscope includes a light source, optical systems configured for generating an excitation light beam and a depletion light beam, optical systems configured for focusing the excitation and depletion light beams on a sample, and systems for collecting and processing data generated by interaction of the excitation and depletion light beams with the sample. Hyperspectral STED data may be analyzed using multivariate curve resolution analysis techniques to deconvolute emission from the multiple fluorophores. The hyperspectral STED microscope described herein can be used for multi-color, subdiffraction imaging of samples (e.g., materials and biological materials) and for analyzing a tissue by Förster Resonance Energy Transfer (“FRET”). |
| Use: Method for multi-color and sub-diffraction imaging of a three-dimensional space, particularly for establishing relationships between the structures in a sample. |
| Advantage: The multi-color and sub-diffraction imaging method involves providing a sample that includes a three-dimensional space having the features of interest, which are smaller than an optical diffraction limit, thus improving the fluorescence microscopy image resolution, multivariate curve resolution analysis of hyperspectral images, spatial resolution of stimulated emission depletion microscopy, reducing the burden, cost and barriers, preventing the back reflection of the light and uncertainty in a fluorescence resonance energy transfer imaging, and accurately measuring fluorophore abundance. |
| Novelty: The multi-color and sub-diffraction imaging method involves providing a sample (136) that includes a three-dimensional space having the features of interest, which are smaller than an optical diffraction limit. The three-dimensional space is labeled with the different fluorophores having a spectrally overlapped excitation and the emission wavelengths. The sample is positioned in a hyperspectral stimulated emission depletion microscope system (100) that is configured to generate an excitation light beam (120) and a depletion light beam (122) coincident with the excitation light beam. |
| Filed: 9/10/2012 |
| Application Number: US13609057A |
| Tech ID: SD 11757.0 |
| This invention was made with Government support under Contract No. DE-NA0003525 awarded by the United States Department of Energy/National Nuclear Security Administration. The Government has certain rights in the invention. |
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