Fluorochrome Panel Selection Using Spectral Stability Metrics
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Solution Overview
Problem
Conventional flow cytometry is limited by a practical limit on the number of fluorochromes that can be used simultaneously due to unavoidable spectral overlap and similarity, restricting panel sizes despite the availability of nearly 100 distinct fluorochrome molecules.
Innovation Solution
A method for identifying a fluorochrome panel by selecting subsets based on a numerical stability metric, such as a condition number, to optimize spectral overlap and increase biological resolution without evaluating antigen data, using a spectral matrix associated with the instrument.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Quantity of substance
If more fluorochromes are used simultaneously in flow cytometry, then the number of detectable markers increases, but spectral overlap increases causing measurement variance and reduced biological resolution
Solution Approach 1:
The patent changes the selection criteria for fluorochrome panels by introducing numerical stability metrics (condition numbers) as a key parameter. Instead of arbitrarily selecting fluorochromes, the system calculates condition numbers for different panel combinations and selects panels with optimal numerical stability, thereby resolving the spectral overlap issue while maintaining high marker throughput
Solution Approach 2:
The patent performs preliminary computational analysis of fluorochrome panel combinations before actual flow cytometry experiments. By pre-calculating condition numbers and spectral overlap characteristics for various panel configurations, the system identifies optimal panels in advance, preventing measurement variance before it occurs
2Adaptability or versatility
If fluorochrome panel size is increased beyond conventional limits, then more biological markers can be characterized, but data variance increases due to spectral similarity
Solution Approach 1:
The patent implements a feedback mechanism where condition numbers calculated from spectral matrices guide the selection and optimization of fluorochrome panels. The system continuously evaluates panel configurations based on their numerical stability metrics and adjusts selections to maintain optimal data quality, enabling larger panels with controlled variance
Solution Approach 2:
The patent makes the fluorochrome panel selection dynamic by allowing the system to evaluate and switch between different panel configurations based on numerical stability requirements. Rather than using fixed panel sizes, the system adaptively determines optimal panel compositions for different experimental conditions while maintaining reliability
3Ease of operation
If conventional fluorochrome selection methods are used, then panel design is simplified, but the number of usable fluorochromes is limited by spectral overlap
Solution Approach 1:
The patent replaces manual, expert-based fluorochrome panel design with an automated computational system. The system uses algorithms to calculate condition numbers and evaluate spectral matrices, automatically identifying optimal panel configurations without requiring manual spectral analysis or expert intuition, thereby enabling larger panels while maintaining ease of use
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 the use of a larger number of fluorochromes in flow cytometry experiments, improving biological resolution and reducing data variance through optimized fluorochrome selection.
Implementation Method 1
the light emitted from fluorescent molecules or fluorescent dye
Implementation Method 2
light at the excitation wavelength scattered by the particle in a narrow angle along a mostly forward direction, referred to as forward-scatter (FSC), the excitation light that is scattered by the particle in an orthogonal direction to the excitation laser, referred to as side-scatter (SSC)
Implementation Method 3
Forward-scattered light, side-scattered light and fluorescent light is detected by photodetectors that are positioned within the particle analyzer
Data Source
AI summary
Methods of identifying a fluorochrome panel suitable for use in a flow cytometric protocol are provided. Methods of interest include receiving an instrument identifier and a request for N fluorochrome identifiers, selecting two or more subsets of N fluorochrome identifiers from a set of fluorochrome identifiers in a spectral matrix associated with the instrument identifier, and identifying the fluorochrome panel from the two or more subsets of N fluorochrome identifiers based on a numerical stability metric calculated for each of the two or more subsets of N fluorochrome identifiers using the spectral signatures associated with the two or more subsets of N fluorochrome identifiers. Systems and non-transitory computer readable storage media for practicing the invention are also provided.