Sheet Illumination Microscope Dynamic Focal Stacking
Find Innovative SolutionsGenerate Solutions
Solution Overview
Problem
Conventional sheet illumination microscopes face challenges in achieving high z resolving power over a wide field-of-view while minimizing the number of acquired images, as a high numerical aperture leads to a narrow illuminated area and increased photobleaching due to prolonged illumination time.
Innovation Solution
A sheet illumination microscope system that moves the condensing position of the light sheet in the illumination direction during the exposure period, combined with a spatial frequency filtering process using a calculation device to enhance the optical transfer characteristic and recover z resolution, allowing for a wider field-of-view with fewer images and reduced photobleaching.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If a high numerical aperture is used to achieve high z resolving power, then the z axis resolution is improved, but the illuminated area becomes narrow and the illumination time increases causing photobleaching
Solution Approach 1:
The illumination optical system dynamically moves the condensing position of the light sheet in the illumination direction during the exposure period. This dynamic adjustment allows the system to maintain a wide field-of-view while preserving high z-axis resolution, and reduces the total illumination time by efficiently capturing the required information across multiple focal planes within a single exposure sequence, thereby minimizing photobleaching.
Solution Approach 2:
The system changes the condensing position parameter of the light sheet during exposure. By varying the focal position of the illumination optical system along the illumination direction, the system can illuminate different depth regions sequentially during the exposure period, achieving high z-resolution through focal stacking while maintaining a wide field-of-view and reducing the need for prolonged illumination.
2Measurement precision
If a high numerical aperture is used to achieve high z resolving power, then the z axis resolution is improved, but the field-of-view becomes narrow
Solution Approach 1:
The illumination optical system dynamically adjusts the condensing position of the light sheet during the exposure period, enabling the system to capture images at multiple focal planes within a single exposure sequence. This dynamic focal stacking approach achieves high z-axis resolution while maintaining a wide field-of-view, as the system can illuminate and capture information from different depth regions without physically moving the sample or detector.
Solution Approach 2:
The system introduces temporal dimension to the illumination process by varying the condensing position during the exposure period. This allows the system to achieve high z-resolution through focal stacking in the time dimension while maintaining a wide field-of-view in the spatial dimensions, effectively adding a temporal degree of freedom to resolve the spatial contradiction.
3Area of stationary object
If the condensing position is moved to expand the illuminated area, then the field-of-view is improved, but the z resolution deteriorates due to expansion of illumination light amount distribution
Solution Approach 1:
The system dynamically moves the condensing position during the exposure period in a controlled sequence, illuminating different focal planes sequentially rather than simultaneously. This dynamic approach allows the system to achieve a wide field-of-view by covering multiple depth regions while maintaining high z-resolution, because each focal plane is illuminated with sufficient intensity during its brief exposure window, and the final image is reconstructed by combining information from all focal planes.
Solution Approach 2:
The system performs preliminary action by pre-planning the sequence of condensing position movements during the exposure period. The illumination optical system is programmed to move through a predetermined sequence of focal planes, ensuring that each region receives appropriate illumination intensity at the right time, thereby achieving both wide field-of-view and high z-resolution without the need for post-acquisition processing to correct illumination non-uniformity.
Data Source
AI summary
An image processing device includes a spatial frequency filtering process unit. The spatial frequency filtering process unit performs a spatial frequency filtering process on image data of a three-dimensional image of the observed object in accordance with the optical transfer characteristic of the microscope apparatus. The spatial frequency filtering process is a process of changing a spatial frequency characteristic included in the three-dimensional image. The three-dimensional image is constructed from a plurality of two-dimensional images of the observed object acquired by the microscope apparatus under a first illumination light amount distribution. The first illumination light amount distribution is formed by moving a condensing position of a light sheet in an illumination direction. The light sheet is emitted to the observed object from the illumination direction.