Fluorescence Image Intensity Mapping to Flow Cytometry Standards

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Solution Overview

Problem

The challenge lies in establishing a mapping relationship between fluorescence intensity representations in image analysis and flow cytometry analysis, making the two methods' results non-comparable and affecting the accuracy of quasi-flow analysis.

Innovation Solution

The method involves performing fluorescence imaging, edge extraction, and segmentation to calculate cumulative, maximum, and average grayscale values, along with bright field diameter, which are then used for flow clustering analysis to align with flow cytometry results.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Productivity

If fluorescence imaging is performed on a large number of to-be-detected targets and image recognition method is used for analysis, then productivity is improved, but measurement precision deteriorates due to inability to establish mapping relationship between different fluorescence intensity representations

Engineering Contradiction:

Improvethroughput of cell analysisVSAvoidaccuracy of fluorescence intensity measurement

Core Design Contradiction:

ProductivityVSMeasurement precision

Solution Approach 1:

The patent transforms image data parameters (pixel coordinates, grayscale values, area) into flow cytometry equivalent parameters (fluorescence intensity, pulse width) through mathematical calculations. Specifically, it calculates cumulative grayscale values, maximum grayscale values, and average grayscale values from segmented image regions, and converts these into fluorescence intensity parameters that match flow cytometry measurement standards, enabling cross-method comparability.

Inventive Principle:

Principle #35Parameter changes

Solution Approach 2:

The patent introduces an intermediary data transformation process that acts as a bridge between image analysis and flow cytometry analysis. This intermediary system performs edge extraction, area segmentation, and grayscale value calculation on fluorescence images, then converts these image-derived parameters into flow cytometry-compatible parameters, allowing results from both methods to be compared and integrated.

Inventive Principle:

Principle #24Intermediary (Mediator)

2Measurement precision

If flow cytometry analysis is used to count fluorescence intensities, then measurement precision is improved, but device complexity increases due to specialized equipment requirements

Engineering Contradiction:

Improveaccuracy of fluorescence intensity measurementVSAvoidcomplexity of detection equipment

Core Design Contradiction:

Measurement precisionVSDevice complexity

Solution Approach 1:

The patent makes the fluorescence microscopic imaging system perform multiple functions: it not only captures fluorescence images for visual analysis but also extracts quantitative fluorescence intensity data through image processing algorithms. This allows the same equipment to serve both imaging and flow-cytometry-like quantification purposes, eliminating the need for separate specialized instruments.

Inventive Principle:

Principle #6Universality (Multi-functionality)

Solution Approach 2:

The patent replaces the physical flow cytometry measurement mechanism (where cells pass through a laser beam and generate voltage pulses) with an image-based measurement mechanism. Instead of using flow cytometry hardware to measure fluorescence intensity, the system uses fluorescence microscopy combined with digital image processing to calculate equivalent parameters from pixel grayscale values, substituting a mechanical measurement system with an optical-digital hybrid system.

Inventive Principle:

Principle #28Mechanics substitution (Replace mechanical system)

3Ease of operation

If fluorescence intensity is represented by pixels and grayscales in image analysis, then ease of operation is improved, but measurement precision deteriorates due to lack of direct comparability with flow cytometry standards

Engineering Contradiction:

Improvesimplicity of image analysisVSAvoidcomparability of fluorescence intensity data

Core Design Contradiction:

Ease of operationVSMeasurement precision

Solution Approach 1:

The patent performs parameter transformation by converting image analysis parameters (pixel coordinates, grayscale values, segmented area) into flow cytometry standard parameters (fluorescence intensity, pulse width). The system calculates cumulative grayscale values from all pixels within segmented cell regions and converts these into fluorescence intensity measurements that directly correspond to flow cytometry data, enabling precise comparison between the two methods.

Inventive Principle:

Principle #35Parameter changes

Data Source

PatentUS11249005B2Method and system for determining fluorescence intensity of fluorescence image

Publication Date: 2022.02.15 SHANGHAI RUIYU BIOTECH

AI summary

A method and a system for fluorescence intensity of a fluorescence image are provided. In the method, fluorescence imaging is performed on a target sample to obtain a fluorescent image. Edge extraction and segmentation is performed on each detection target in the fluorescence image, to obtain the fluorescent image area of each detection target in the fluorescence image. At least one of a cumulative gray-scale value, a maximum gray-scale value and an average gray-scale value of the fluorescence image region and a diameter value of a bright field image region of each detected target is calculated. Then, the flow clustering analysis is calculated based on at least one of the cumulative grayscale value, the maximum grayscale value, the average grayscale value, and the bright field diameter value.