Super-Resolution Microscopy Using Multi-Frequency Pulsed Laser Segmentation
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Solution Overview
Problem
Conventional unstained microscopy techniques face challenges in achieving high optical resolution and discrimination capability for observing samples without staining, particularly in satisfying the requirements of plane resolution below 50 nm and depth resolution below 100 nm.
Innovation Solution
A super-resolution observation device and method utilizing a combination of pulsed laser lights with different optical frequencies, where a first light with frequency ω1 excites the sample, a second light with frequency ω2′ induces stimulated emission in an overlapping region, and a third light with frequency ω2 generates signal light in a non-overlapping region, allowing for signal extraction beyond the diffraction limit.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If conventional unstained microscopy is used to observe samples without staining, then the sample remains in an intact state, but the optical resolution and discrimination capability are insufficient to achieve plane resolution below 50 nm and depth resolution below 100 nm
Solution Approach 1:
The patent segments the observation process into distinct temporal phases using multiple pulsed laser lights with different frequencies. The first pulsed light (frequency ω1) excites the sample, the second pulsed light (frequency ω2') induces stimulated emission in the overlap region, and the third pulsed light (frequency ω2) generates signal light in the non-overlap region. This temporal and spatial segmentation enables super-resolution by isolating the detection origin to a region smaller than the diffraction limit, thereby improving optical resolution while maintaining sample integrity without staining
Solution Approach 2:
The patent employs periodic pulsed laser illumination with different frequencies in a sequence to achieve super-resolution. The periodic application of excitation light, stimulated emission light, and signal-generating light allows for repeated measurement cycles, enabling high-resolution observation through time-resolved detection while keeping the sample in an intact unstained state
2Measurement precision
If multiple pulsed laser lights with different frequencies are used to achieve super-resolution, then spatial resolution is enhanced, but the device complexity increases
Solution Approach 1:
The patent combines multiple pulsed laser lights with different frequencies (ω1, ω2', ω2) into a unified illumination system that shares common optical components such as the objective lens and detection path. By merging the excitation, stimulated emission, and signal-generating functions into a coordinated multi-frequency pulsed system, the patent achieves super-resolution while reducing the overall device complexity compared to using completely separate independent systems for each function
Solution Approach 2:
The illumination optical system is designed with multi-functionality, where a single system performs multiple roles: exciting the sample, inducing stimulated emission, and generating signal light through the use of multiple pulsed laser lights with different frequencies. This universal design approach allows one system to achieve super-resolution capabilities without requiring separate specialized devices for each function, thereby managing device complexity
3Measurement precision
If the detection origin is limited to a non-overlapping region smaller than the resolution limit, then discrimination capability is improved, but the area available for observation is reduced
Solution Approach 1:
The patent transitions from spatial resolution limits to temporal resolution by using time-resolved detection of signal light generated at different frequencies. By observing the sample in the time domain through sequential pulsed illumination and detecting the temporal characteristics of emitted light, the system achieves high discrimination capability in a limited non-overlap region while effectively expanding the observable information through time-dimensional analysis
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 super-resolution observation of samples without staining by limiting the detection origin to a non-overlapping region smaller than the resolution limit, enhancing spatial resolution and discrimination capability.
Implementation Method 1
collecting a first illuminating light having a first optical frequency ω1 on a first region of an observation object
Implementation Method 2
collecting a second illuminating light having a second optical frequency ω2' on a second region partially overlapping the first region
Implementation Method 3
collecting a third illuminating light having a third optical frequency ω2 on a third region containing a non-overlap region
Implementation Method 4
an extraction unit extracting a signal light generated in accordance with a change in an energy level of a substance in the non-overlap region from a light generated in all of the first region, the second region, and the third region
Data Source
AI summary
A super-resolution observation device includes an illumination optical system collecting a first illuminating light having a first optical frequency ω1 on a first region of an observation object, collecting a second illuminating light having a second optical frequency ω2′ on a second region partially overlapping the first region, and collecting a third illuminating light having a third optical frequency ω2 on a third region containing a non-overlap region which is a region of the first region and does not overlap the second region; and an extraction unit extracting a signal light generated in accordance with a change in an energy level of a substance in the non-overlap region from a light generated in all of the first region, the second region, and the third region.