Confocal Microscope Scanning for Moving Particle Detection
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Solution Overview
Problem
Current optical microscopic techniques struggle to effectively image light-emitting particles in dynamic conditions within thick samples, as raster scan modes fail to capture moving particles accurately, and evanescent light microscopes are limited to surface observations, making it difficult to detect and track particles in depth.
Innovation Solution
An optical microscope system using a confocal or multiphoton microscope with a light detection region that moves multiple times within divided subregions, generating time series light intensity data to individually detect and track light-emitting particles, allowing for precise positioning and imaging of particles in three-dimensional spaces.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Area of stationary object
If raster scan mode is used to image light-emitting particles, then the imaging process can cover a large region, but moving particles cannot be accurately captured because their position changes during multiple scans
Solution Approach 1:
The imaging region is divided into multiple subregions, and the light detection region performs multiple continuous scans within each subregion before moving to the next subregion. This segmentation allows sufficient time to capture moving particles within each small subregion while maintaining coverage of the entire imaging region through systematic progression across all subregions.
2Measurement precision
If evanescent light microscope is used, then background light is reduced and single molecule level detection is possible, but observation is limited to surface region only
Solution Approach 1:
The light detection region is made dynamically movable through continuous scanning within each subregion. This dynamic scanning approach allows the detection region to systematically explore deeper regions of the sample over time, overcoming the static surface limitation of evanescent light microscopes while maintaining high detection sensitivity through multiple passes.
3Illumination intensity
If multiple scans are performed in raster mode, then light intensity can be integrated for better signal, but particles that move during scanning cannot be effectively integrated
Solution Approach 1:
The scanning process is segmented into multiple continuous scans within each subregion. By confining multiple scans to small subregions rather than scanning the entire region repeatedly, particles are more likely to remain within the same subregion throughout the multiple scans, enabling effective signal integration while maintaining spatial accuracy.
Applied Scientific Principles
This section explains which scientific principles are used to turn an abstract innovation direction into a practical engineering solution.
Function Achieved in This Case
This method enables accurate detection and imaging of dynamically moving light-emitting particles in thick samples, providing detailed two- or three-dimensional images and improving the detection efficiency of particles, even when they change position rapidly.
Implementation Method 1
the light emitted from a light-emitting particle may be fluorescence
Implementation Method 2
the light emitted from a light-emitting particle may be phosphorescence
Implementation Method 3
the light emitted from a light-emitting particle may be chemoluminescence
Implementation Method 4
the light emitted from a light-emitting particle may be bioluminescence
Implementation Method 5
the light emitted from a light-emitting particle may be scattering light
Implementation Method 6
when the light detection region encompasses a light-emitting particle distributed and moving at random in the sample solution
Data Source
AI summary
There is provided a microscopic observation technique capable of detecting a light-emitting object or a light-emitting particle moving in a thick sample by the scanning molecule counting method. In the inventive technique, the light from a light detection region of is detected the optical system of a confocal or multiphoton microscope is detected with while moving the light detection region in each observed subregion obtained by dividing a region to be observed into plural regions; the signal of the light from a light-emitting particle is individually detected; and the position of the light-emitting particle corresponding to the detected signal is determined in the region to be observed. The moving of the position of the light detection region in each observed subregion is performed continuously in at least two directions or and/or continuously multiple times in each observed subregion.