3D Microscope Focusing via Phase Intensity Analysis
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Solution Overview
Problem
Current microscopes face challenges in maintaining precise three-dimensional focusing due to positional drift caused by environmental vibrations and temperature variations, limiting the accuracy and stability of imaging, especially in ultra-resolved optical microscopes like PALM or STORM, where existing methods offer precision up to 50 nm along the optical axis and require complex setups or feedback control systems.
Innovation Solution
A device and method utilizing a detector to analyze the phase and intensity of an electromagnetic field interacting with a reference object of varying refractive index, integrated with a microscope, allowing for simultaneous measurement of axial and lateral position variations, enabling three-dimensional focusing with a resolution below 10 nm without the need for complex grating fabrication or feedback control.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If conventional focusing methods (IR LED reflection) are used, then the device complexity remains low, but the measurement precision along the optical axis does not exceed 50 nm
Solution Approach 1:
The patent introduces a reference object with a known complex refractive index as an intermediary element between the light source and detector. This reference object interacts with the incident light wave to produce characteristic phase and intensity patterns that enable precise measurement of axial position variations, achieving sub-10 nm precision without complex feedback control systems
Solution Approach 2:
The patent utilizes changes in optical parameters (phase and intensity) of the electromagnetic field as the reference object moves along the optical axis. By detecting these parameter variations and comparing them with pre-calculated patterns, the system achieves high-precision focusing measurement through parameter analysis rather than mechanical complexity
2Measurement precision
If micrometric markers with transverse dimension measurement are used, then the measurement precision reaches about twenty nanometers, but a closed-loop feedback control system is required
Solution Approach 1:
The reference object serves as a mediator that converts axial position information into characteristic phase and intensity patterns detectable in a single measurement plane. This eliminates the need for transverse dimension measurements and closed-loop feedback, as the axial position can be directly determined from the optical field characteristics
Solution Approach 2:
The patent replaces mechanical feedback control systems with an optical field analysis approach. By analyzing the phase and intensity distribution of the electromagnetic field interacting with the reference object, the system determines axial position variations optically rather than through mechanical measurement and feedback loops
3Measurement precision
If 2-dimensional microscopic grating is used, then theoretical accuracies of a few tens of nanometers are achieved, but complex grating fabrication is required
Solution Approach 1:
The patent extracts the essential function of the grating (providing reference patterns for measurement) and implements it through a simple reference object with known refractive index properties. This eliminates the need for complex 2D grating fabrication while maintaining measurement accuracy, as the reference object's optical interaction naturally produces the necessary phase and intensity patterns
Solution Approach 2:
Instead of relying on geometric structures like gratings, the patent uses changes in optical parameters (phase and intensity) resulting from the interaction between light and the reference object's complex refractive index. This approach achieves high precision through optical property variations rather than complex geometric fabrication
4Measurement precision
If data acquisition is performed with PALM microscope, then ultra-resolved imaging is achieved, but data acquisition time becomes long (several hours), increasing stability requirements
Solution Approach 1:
The patent performs preliminary calibration by calculating the expected phase and intensity patterns for known axial positions of the reference object. During actual measurement, this pre-calculated information enables rapid determination of position drifts without requiring long acquisition times, thus maintaining ultra-resolution while reducing measurement duration
Solution Approach 2:
The patent replaces time-consuming iterative focusing procedures with a rapid optical field analysis method. By analyzing the phase and intensity distribution in a single measurement plane and comparing with pre-calculated patterns, the system achieves both ultra-resolution and fast acquisition, eliminating the several-hour data collection requirement
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 solution provides precise three-dimensional focusing with a resolution of less than 10 nm, enhancing the stability and accuracy of imaging by determining the drifts of the microscope, allowing for automatic focusing and compensating for environmental disturbances, and can be easily integrated into conventional microscopes.
Implementation Method 1
the reference object exhibiting a complex refractive index whose real part and/or imaginary part is different respectively from the real part and/or from the imaginary part of the refractive index of the reference medium
Implementation Method 2
an image in phase and in intensity of an electromagnetic field resulting from the interaction of an incident light wave with a reference object
Data Source
Figure 1A
Figure 1B
Figure 2A~2C
AI summary
The invention relates to a device (II) for three-dimensional focusing in a microscope (I) with a given resolution, comprising: - a detector (119) making it possible to determine, in a given measuring plane, a phase and intensity image of an electromagnetic field resulting from the interaction of an incident light wave with a reference object (105) situated in a reference medium with a given index and having a complex refractive index, the real part and/or the imaginary part of which differs from the real part and/or the imaginary part, respectively, of the refractive index of the reference medium; - computation means (121) making it possible to determine a three-dimensional position variation of the reference object (105) from at least one phase and intensity image.