Phase Distribution Measurement for Dye-Free 3D Cell Observation
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Solution Overview
Problem
Current methods for observing three-dimensional structures of living cells and tissues, such as those using fluorescence confocal microscopes, require fluorochromes or fluorescent proteins, which pose risks and are not suitable for clinical applications, especially in regenerative medicine where mutation and alteration detection is crucial.
Innovation Solution
A phase distribution measurement method and apparatus that converts phase distribution into image intensity distribution using a microscope, changing image contrast to form multiple images, calculating and normalizing phase components, and performing deconvolution processes to obtain accurate phase distributions without the need for dyes or fluorochromes, allowing for precise observation of biological samples.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If fluorescence confocal microscope is used to observe three-dimensional structure of living cells and tissues, then good three-dimensional resolution is achieved, but fluorochromes or fluorescent proteins are required which pose risks of cellular damage and mutation
Solution Approach 1:
The invention extracts and eliminates the harmful element (fluorochromes and fluorescent proteins) from the observation system while preserving the useful function (three-dimensional structure observation). By using phase contrast microscopy combined with image processing, the method achieves 3D observation without requiring any exogenous fluorescent labels that could damage or mutate cells.
Solution Approach 2:
The invention replaces the optical-mechanical fluorescence excitation system with a phase contrast optical system combined with computational image processing. Instead of using fluorescent excitation light that requires dye molecules, the system uses phase contrast microscopy to capture optical path difference information and reconstructs 3D structures through digital deconvolution and integration processes.
2Object-affected harmful factors
If phase contrast microscopy is used to observe living cells without dyes, then cellular damage is reduced, but the ability to detect mutations and alterations is insufficient compared to fluorescence methods
Solution Approach 1:
The invention performs preliminary actions of capturing multiple phase contrast images at different focal planes and processing them through deconvolution and integration algorithms before final observation. By pre-processing the optical data to extract quantitative phase information and reconstruct 3D phase distributions, the system enhances its ability to detect subtle cellular alterations without requiring fluorescent labeling.
Solution Approach 2:
The invention introduces computational image processing algorithms as an intermediary between the phase contrast microscope and the final observation result. This intermediary layer processes the raw optical data to extract quantitative phase information, perform deconvolution to remove optical aberrations, and integrate images from multiple focal planes, thereby enhancing the detection capability for mutations and alterations while maintaining the dye-free advantage.
3Measurement precision
If multiple images with different contrasts are captured and processed through deconvolution, then accurate phase distribution is obtained, but the observation process becomes more complex
Solution Approach 1:
The invention creates a universal image processing framework that handles multiple image contrasts and focal planes through a single integrated deconvolution and integration algorithm. This multi-functional processing system can accommodate various input conditions (different contrasts, different focal planes) and produces consistent quantitative phase distribution results, thereby managing complexity through algorithmic unification rather than separate processing procedures.
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
Enables accurate, dye-free observation of three-dimensional structures, reducing the risk of cellular damage and mutation, and facilitating the detection of mutations and alterations in living cells and tissues, particularly in regenerative medicine applications.
Implementation Method 1
a microscope that converts phase distribution into image intensity distribution
Implementation Method 2
calculating phase distribution of a refraction component formed by light refracted inside the biological sample
Implementation Method 3
calculating phase distribution of a structure component formed by light diffracted in a structure inside the biological sample
Data Source
AI summary
A phase distribution measurement method inside a biological sample includes taking in an optical image of the sample formed by a microscope to form a plurality of images with different image contrasts; calculating a component corresponding to phase distribution of the sample and a component corresponding to other than the phase distribution, and dividing the component corresponding to the phase distribution by the component corresponding to other than the phase distribution to forma normalized phase component image; breaking down the phase component image into a plurality of frequency components; performing a deconvolution process to each of the frequency components using an optical response character corresponding to each, and calculating phase distribution of a refraction component and phase distribution of a structure component; and calculating phase distribution of the sample by compounding the phase distribution of the refraction component and the phase distribution of the structure component.