Fluorescence Imaging Control Using Control Markers for Biological Samples
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Solution Overview
Problem
The existing methods for producing fluorescence images of biological samples stained with multiple fluorescent materials face challenges in focusing without accelerating the discoloration of the fluorescent materials or overloading the sample, especially when connecting enlarged sections of the sample, which leads to reduced microscopic visibility due to degradation and discoloration over time, particularly when samples are transported and diagnosed in different facilities.
Innovation Solution
A method and apparatus that utilize a first and second fluorescent material with different excitation wavelengths, where the image is focused and captured using a data processing unit that controls the excitation light sources, allowing for dark field and bright field imaging modes to adjust focus without exciting the fluorescent material intended for the target, thereby preventing discoloration and overloading the sample.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Area of stationary object
If the biological sample is sectioned and enlargements of sectioned parts are connected together to obtain an image of the entire sample, then the entire image can be formed on the imaging surface, but it takes a longer time to connect the enlargements of the parts due to moving the stage to focus on each part
Solution Approach 1:
The patent applies preliminary action by capturing an image of the control marker before the fluorescence staining is completed. This allows the focusing position to be determined in advance based on the control marker image, eliminating the need to move the stage to focus on each part during the connection process. The control marker image serves as a pre-prepared reference that enables rapid positioning without time-consuming stage movements.
2Area of stationary object
If the stage is moved to focus on each part of the sample to connect enlargements, then the entire image can be formed, but the fluorescent material discoloration is accelerated and the biological sample is overloaded
Solution Approach 1:
The patent uses preliminary action by capturing the control marker image before fluorescence staining is completed. This allows the focusing position to be determined in advance, eliminating the need for repeated stage movements during the connection process. By determining the focus position beforehand based on the control marker, the patent avoids repeated exposure to excitation light that would cause fluorescent material discoloration and sample overloading.
Solution Approach 2:
The patent introduces the control marker as an intermediary substance that enables focusing without directly exciting the fluorescent material. The control marker serves as a mediator to determine the focusing position, allowing the system to focus on the sample area without repeatedly exposing the fluorescent material to excitation light, thus preventing discoloration and overloading.
3Loss of information
If the biological sample is stained with fluorescence staining to determine malignant tumor characteristics, then molecular biological information can be obtained, but the sample becomes clear and colorless in unexcited state making it difficult to focus on the image
Solution Approach 1:
The patent introduces the control marker as an intermediary substance that provides contrast in the unexcited state. The control marker absorbs excitation light and produces fluorescence that can be detected in the unexcited state, creating sufficient contrast to focus on the image. This intermediary allows the system to focus on the sample area without requiring the fluorescent material to be in an excited state, thus preserving the molecular biological information while enabling image detection.
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 approach enables accurate focusing on biological samples without accelerating the discoloration of the fluorescent material or overloading the sample, maintaining image clarity and reducing the time required for image capture, thus addressing the issues of sample degradation and visibility during transportation and diagnosis.
Implementation Method 1
a first fluorescent material and a second fluorescent material having different excitation wavelengths; a first light source configured to irradiate the biological sample thereby allowing focusing of the biological sample based on fluorescence of the first fluorescent material; and a second light source configured to irradiate the biological sample thereby allowing capturing of the image based on fluorescence of the second fluorescent material
Data Source
AI summary
Method and apparatus for producing an image associated with a biological sample is disclosed. A processor focuses on a portion of the biological sample based on fluorescence of a first fluorescent material and captures an image of the portion of the biological sample based on fluorescence of the first fluorescent material and the second fluorescent material.