Three-Dimensional FISH Analysis for Genetically Abnormal Cell Detection
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Solution Overview
Problem
Current methods for analyzing circulating tumor cells (CTCs) using fluorescence in situ hybridization (FISH) face challenges in detecting CTCs without specific surface markers, leading to exclusion of marker-negative cells and difficulty in accurately distinguishing between overlapping fluorescence signals in two-dimensional images, resulting in reduced detection accuracy.
Innovation Solution
A method involving immunostaining against cell surface antigens and subsequent three-dimensional image analysis of fluorescence signals using nucleic acid probes specific to cancer genes, allowing for the detection and analysis of genetically abnormal cells, including CTCs, by separating and removing non-genetically abnormal cells and using antibodies to distinguish between cell groups.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If conventional two-dimensional FISH analysis is used to detect CTCs, then the detection process is simple, but the detection accuracy is reduced due to difficulty in distinguishing overlapping fluorescence signals
Solution Approach 1:
The patent transitions from two-dimensional FISH signal analysis to three-dimensional image analysis. By reconstructing three-dimensional images from multiple focal planes and analyzing fluorescence signal distribution in three-dimensional space, the method enables accurate distinction between overlapping signals that cannot be resolved in two dimensions, thereby improving detection accuracy without excessive complexity increase
2Productivity
If immunostaining against cell surface antigens is performed to identify CTCs, then cells can be separated and analyzed, but marker-negative CTCs are excluded from detection
Solution Approach 1:
The patent combines immunostaining against cell surface antigens with FISH analysis in a sequential workflow. Cells are first immunostained to identify and separate CTCs based on surface markers, then subjected to FISH analysis to detect genetic abnormalities. This combination maintains high detection efficiency through positive selection while adding comprehensive genetic analysis capability, addressing both productivity and adaptability requirements
3Measurement precision
If FISH is performed on all cells in blood samples, then no marker-negative cells are excluded, but the large amount of leukocytes interferes with CTC detection
Solution Approach 1:
The patent extracts and isolates CTCs from blood samples using immunostaining against epithelial cell surface antigens before performing FISH analysis. This extraction step separates the target CTCs from the large quantity of non-target leukocytes, enabling focused analysis on relevant cells and improving detection accuracy while reducing unnecessary processing of irrelevant cells
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 highly accurate and simple detection of genetically abnormal cells, even in trace amounts, by clearly distinguishing between immunostaining and FISH signals in three-dimensional images, improving the analysis of CTCs and reducing the risk of excluding marker-negative cells.
Implementation Method 1
immunostaining of cells contained in the cell sample using one or more antibodies against a cell surface antigen of genetically abnormal cells
Implementation Method 2
conducting FISH (fluorescence in situ hybridization) to the cells contained in the cell sample using one or more nucleic acid probes specifically binding to DNA of a cancer gene
Implementation Method 3
analyzing fluorescence signals from the nucleic acid probes in the cells contained in the cell sample
Data Source
Figure 1(A)~1(E)
Figure 2(A)~2(E)
Figure 3(A)~3(B)
AI summary
The present invention provides a method for detecting and analyzing cells having gene abnormality in a highly accurate and simple manner using FISH. The present invention relates to a method for analyzing genetically abnormal cells, the method including the steps of (a) preparing a cell sample containing eukaryotic cells, (b) conducting an antigen-antibody reaction to cells contained in the cell sample using one or more antibodies which specifically bind to a molecule existing on the cell surface of eukaryotic cells after the step (a), (c) subjecting the cells contained in the cell sample to a permeation treatment after the step (b), (d) subjecting the cells contained in the cell sample to an immobilization treatment after the step (b), (e) conducting FISH to the cells contained in the cell sample using one or more nucleic acid probes after the step (d), (f) analyzing fluorescence signals from the nucleic acid probes in the cells contained in the cell sample using a three-dimensional image analysis method after the step (e), and (g) determining whether the cells contained in the cell sample are genetically abnormal cells or not based on the results of the step (b) and the results of the step (f).