Cell Identification Using Fluorescence and White Light Imaging
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Solution Overview
Problem
Current methods for identifying rare cells in blood samples, such as circulating tumor cells and fetal nucleated red blood cells, face challenges due to low target-to-background cell ratios and suboptimal antibody binding accuracy, leading to inadequate identification and sorting efficiency.
Innovation Solution
A cell identification method utilizing fluorescent markers and dual imaging techniques, including fluorescence and white light scans, to accurately mark and differentiate target cells from non-target cells based on specific fluorescence signal brightness and cell morphology, enhancing the identification of rare cells.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If blood smears are used to identify target cells under the microscope via chemical staining, then the identification process is simple and inexpensive, but the identification accuracy is insufficient due to low target-to-background cell ratios (1:10^7 to 10^9)
Solution Approach 1:
The identification process is divided into multiple sequential steps: initial fluorescence screening to identify candidate cells, followed by white light imaging for morphological verification. This segmentation allows the system to first filter the vast majority of background cells using fluorescence markers, then apply more complex morphological analysis only to the small subset of candidate cells, thereby maintaining high accuracy while managing complexity
Solution Approach 2:
Fluorescence markers serve as an intermediary that enables selective visualization of target cells against the background. By introducing fluorescently-labeled antibodies that specifically bind to target cell surface markers, the system converts the undetectable rare cells into visible fluorescent signals, dramatically improving the effective target-to-background ratio from 1:10^7-10^9 to a much higher ratio that enables accurate identification
2Measurement precision
If specific antibodies are used to mark target cells, then target cell identification is enhanced, but the binding rate and accuracy do not meet the minimum standard (99.5%) for clinical identification
Solution Approach 1:
The system performs preliminary fluorescence-based screening to identify candidate target cells before applying morphological analysis. By pre-selecting cells that exhibit the expected fluorescence signal pattern, the system ensures that subsequent morphological verification is applied only to relevant candidates, thereby improving overall identification reliability and ensuring that the final accuracy meets the 99.5% clinical standard
Solution Approach 2:
The system uses a feedback mechanism where fluorescence signal intensity and distribution patterns are analyzed to confirm proper antibody binding. Cells that do not exhibit the expected fluorescence characteristics are excluded from further analysis, while those that do are subjected to morphological verification. This feedback loop ensures that only properly bound and correctly identified cells are counted, maintaining reliability above 99.5%
3Measurement precision
If multiple fluorescent markers are used to mark different cell components, then cell identification accuracy is improved, but the device complexity and analysis time increase
Solution Approach 1:
The analysis is segmented into two distinct phases: a rapid fluorescence screening phase that uses multiple markers to identify candidate cells, followed by a targeted white light imaging phase that performs detailed morphological analysis only on those candidates. This segmentation reduces total analysis time compared to performing complete morphological analysis on all cells, while maintaining high accuracy through the use of multiple fluorescent markers for initial identification
Solution Approach 2:
The system applies partial action by using multiple fluorescent markers to screen and identify candidate cells, then applying the more time-consuming white light morphological analysis only to a small subset of those candidates rather than to all cells in the sample. This partial application of the full analysis protocol to only necessary cases significantly reduces total analysis time while maintaining identification 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 significantly improves the accuracy of rare cell identification by using fluorescent markers like Hoechst 33342, anti-EpCAM, and anti-CD45 antibodies, and white light imaging to correctly identify target cells with high precision, even at low concentrations, thereby overcoming previous limitations in clinical identification.
Implementation Method 1
fluorescent markers include a first fluorescent marker for marking nuclei, a second fluorescent marker for marking the target cells, and a third fluorescent marker for marking the target cells or the non-target cells. A fluorescence scan is performed on the analysis sample respectively with a first fluorescence waveband corresponding to the first fluorescent marker, a second fluorescence waveband corresponding to the second fluorescent marker, and a third fluorescence waveband corresponding to the third fluorescent marker
Data Source
AI summary
Provided are a cell identification method for identifying target cells by combining the results of fluorescence intensity analysis, fluorescence image analysis, and white light image analysis.