Fluorescence Signal Separation from Autofluorescence Using Spectral Unmixing

Solution Overview

Problem

Conventional immunofluorescence staining methods face limitations in visualizing antigens due to restricted types of fluorescent pigments that can be coupled to primary antibodies, leading to buried fluorescence signals within autofluorescence signals from specimens, making detection difficult.

Innovation Solution

An information processing apparatus and method that separates fluorescence signals from autofluorescence signals in image information using specimen and reagent information, allowing for accurate extraction and analysis of fluorescence signals without needing unlabeled specimen measurements.

Engineering Contradictions & Design Principles

VSEngineering Contradiction Analysis

1Adaptability or versatility

If a fluorescent reagent is used to visualize antigens, then the types of visualizable antigens are limited by the types of fluorescent pigments that can be coupled to primary antibodies, but the fluorescence signal can be detected

Engineering Contradiction:

Core Design Contradiction:

Adaptability or versatilityVSMeasurement precision

Solution Approach 1:

The patent segments the fluorescence signal into two distinct components: autofluorescence signal from the specimen and specific fluorescence signal from the fluorescent reagent. By separating these signals through spectral analysis, the system enables detection of fluorescent reagents even when their emission spectra overlap with autofluorescence, thus resolving the contradiction between signal detectability and antigen type versatility

Inventive Principle:

Principle #1Segmentation

Solution Approach 2:

The patent introduces an information processing apparatus as an intermediary that performs spectral unmixing between the raw fluorescence image and the final separated signals. This intermediary system uses reference spectra of autofluorescence and fluorescent reagents to mathematically decompose the mixed signal, enabling accurate detection of the specific fluorescence signal even when buried in autofluorescence

Inventive Principle:

Principle #24Intermediary (Mediator)

2Measurement precision

If conventional immunofluorescence staining is used, then the fluorescence signal can be detected, but the signal is buried in autofluorescence signal making detection difficult

Engineering Contradiction:

Core Design Contradiction:

Measurement precisionVSObject-generated harmful factors

Solution Approach 1:

The patent extracts the specific fluorescence signal from the mixed fluorescence image by removing the autofluorescence component. Through spectral unmixing using reference spectra, the system mathematically separates and extracts the signal of interest from the harmful autofluorescence background, enabling clear detection of the fluorescence signal even when initially buried

Inventive Principle:

Principle #2Taking out (Extraction)

Solution Approach 2:

The patent converts the harmful autofluorescence signal into a useful component for signal separation. By measuring and utilizing the autofluorescence reference spectrum, the system uses the previously harmful background signal as a reference to identify and remove itself, thereby converting the harm into a benefit for achieving accurate signal detection

Inventive Principle:

Principle #22Blessing in disguise (Convert harm into benefit)

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 precise separation and detection of fluorescence signals from autofluorescence signals, reducing practitioner burden and improving signal accuracy by using specimen and reagent information to differentiate between the two signals.

Implementation Method 1

separating a fluorescence signal of a fluorescent reagent from an autofluorescence signal of a specimen based on information regarding a spectrum of a fluorescence component included in the fluorescent reagent and information regarding an autofluorescence component included in the specimen

Methodology Applied

Scientific EffectFluorescence: Fluorescence

Data Source

PatentEP3827248B1Information processing apparatus and microscope for separating the fluorescence of a fluorescent reageant from the autofluorescence of a specimen

Publication Date: 2024.06.12

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AI summary

Provided is an information processing apparatus including an image acquisition section (112), an information acquisition section (111), and an analysis section (131). The image acquisition section (112) acquires image information regarding a specimen stained by a fluorescent reagent (30). The information acquisition section (111) acquires information regarding the specimen (11) and information regarding the fluorescent reagent (10). The analysis section (131) separates a first fluorescence signal and a second fluorescence signal from the image information in accordance with the information regarding the specimen and the information regarding the fluorescent reagent.