Spectral Observation System Tunable Laser Masking Shield
Solution Overview
Problem
In spectral imaging of samples with multiple fluorescent reagents, necessary data can be lost due to exclusion of specific wavelengths from the detection range, particularly when the exciting wavelengths of different fluorescent reagents overlap within each other's emission spectra.
Innovation Solution
A spectral observation system and method that employs a confocal microscope with a tunable laser source, adjustable gratings, and a masking shield to selectively exclude specific channels from the detection range, allowing for sequential imaging of each fluorescent reagent's emission spectrum without data loss by switching the detection range and laser wavelengths to capture complete spectral data.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Reliability
If a masking shield is disposed to exclude the channel corresponding to the exciting wavelength from the detection wavelength range, then the fluorescence detection is improved by preventing excitation light interference, but the necessary spectral data may be lost when the exciting wavelength overlaps with the emission spectrum of another fluorescent reagent
Solution Approach 1:
The masking shield is made movable rather than fixed, allowing it to be dynamically adjusted to different positions. This enables the system to selectively exclude specific wavelength channels from detection based on the particular fluorescent reagents being observed, while still capturing complete spectral data for analysis. The movable mechanism allows flexible adaptation to different experimental conditions without permanent loss of detection capability.
Solution Approach 2:
The system changes the detection wavelength range parameters by adjusting the masking shield position according to different fluorescent reagent combinations. By dynamically modifying which wavelength channels are excluded from detection, the system optimizes the balance between preventing excitation light interference and capturing complete emission spectra for accurate spectral unmixing and quantitative analysis.
2Productivity
If multiple fluorescent reagents are excited simultaneously for spectral imaging, then the observation efficiency is improved, but the spectral data may contain interference from overlapping excitation and emission wavelengths
Solution Approach 1:
The system uses spectral unmixing algorithms that analyze the complete spectral data collected from simultaneous multi-color fluorescence imaging. By incorporating feedback from the full spectral information (including wavelengths that would normally be excluded), the system can mathematically separate and quantify the contribution of each fluorescent reagent, thereby maintaining high imaging efficiency while achieving accurate measurement despite spectral overlaps.
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 acquisition of comprehensive spectral data for each fluorescent reagent without gaps, even when their exciting wavelengths overlap, ensuring accurate contribution analysis and minimizing interference-induced fading.
Implementation Method 1
A spectral imaging fluorescent laser microscope irradiates excitation light to the sample, performs a spectral diffraction of fluorescence generated there
Implementation Method 2
performs a spectral diffraction of fluorescence generated there by a grating (diffraction grating) and so on, and detects intensity of each wavelength component
Data Source
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AI summary
A proposition of the present invention is to acquire necessary data without any chasm in an observation of a specimen containing plural kinds of substances of which exciting wavelengths are different. Accordingly, a spectral observation method of the present invention, irradiating light to the specimen containing plural kinds of substances (RFP, GFP, and so on) of which exciting wavelengths are different, and detecting spectra of light emitted from the specimen, the spectral observation method sequentially acquires spectral data of substances from the one of which exciting wavelength is long while switching a wavelength of the irradiated light among respective exciting wavelengths of the plural kinds of substances (S13, S15), and excludes the exciting wavelength of the substance corresponding to the spectral data to be acquired from detection wavelengths of the spectra each when acquiring the spectral data of the spectral data of the respective substances (S12).