Optical Analysis Using Intensity Ratios for Single Particle Identification
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Solution Overview
Problem
Current optical analysis techniques using confocal microscopes struggle to characterize and identify light-emitting particles at concentrations below 1 nM, as the absolute light intensity values are not unique to individual particles due to varying excitation light intensity across the detection region, making it difficult to distinguish between different types and sizes of particles, especially in cases of weak interactions or low concentrations.
Innovation Solution
The method involves using particles with at least two light-emitting sites of different wavelengths, where the ratio of light intensities from these sites remains constant regardless of the detection region position, allowing for characterization and identification based on this invariant ratio, enabling the discrimination of different types and sizes of particles even at low concentrations.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If the absolute light intensity value is used to characterize light-emitting particles, then the measurement is simple, but the identification becomes inaccurate at low concentrations due to varying excitation light intensity across the detection region
Solution Approach 1:
The patent transforms the measurement parameter from absolute light intensity to the ratio of light intensities at two different wavelengths. This parameter transformation eliminates the influence of excitation light intensity variations across the detection region, enabling accurate particle identification even at low concentrations where particles may be located at different positions within the detection volume.
2Quantity of substance
If the concentration of light-emitting particles is reduced to detect rare or expensive samples, then the sample amount is minimized, but the detection sensitivity decreases making it difficult to distinguish individual particles
Solution Approach 1:
The patent utilizes the emission characteristics at two different wavelengths (colors) to identify and characterize individual particles. By measuring the intensity ratio at these two wavelengths, the system can distinguish between different types of particles and maintain detection sensitivity even when the total number of particles in the detection region is very low, enabling analysis of rare or expensive samples.
3Productivity
If the detection region is enlarged to increase the probability of detecting particles, then the detection efficiency is improved, but the spatial resolution decreases making it difficult to obtain precise localization information
Solution Approach 1:
The patent maintains a small detection region (confocal volume) to preserve spatial resolution and precise localization information. By measuring the intensity ratio at two wavelengths rather than relying on absolute intensity, the system achieves sufficient detection sensitivity even with the limited number of particles that pass through the small confocal volume, thus resolving the contradiction between detection efficiency and spatial resolution.
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 approach allows for the accurate characterization and identification of light-emitting particles at concentrations lower than previously possible, expanding the applicability of scanning molecule counting methods to detect and analyze particles in solutions with extremely low concentrations and weak interactions.
Implementation Method 1
the light emitted from a light-emitting particle may be fluorescence, phosphorescence, chemiluminescence, bioluminescence, scattered light, etc.
Implementation Method 2
the light emitted from a light-emitting particle may be fluorescence, phosphorescence, chemiluminescence, bioluminescence, scattered light, etc.
Data Source
AI summary
There is provided a scanning molecule counting method using an optical measurement with a confocal microscope or a multiphoton microscope, enabling characterization of a light-emitting particle or identification of a light-emitting particle with emitted light intensity of a single light-emitting particle measured individually. In the inventive optical analysis technique, with reference to the ratio of the intensities of simultaneously generated signals of the lights of at least two light-emitting sites having mutually different emission wavelengths, possessed by a light-emitting particle contained in a sample solution, the intensities being measured with moving the position of the light detection region of an optical system by changing the optical path of the optical system, a single light-emitting particle corresponding to the signals is identified, and the kind, the size, etc. of the light-emitting particle is identified.