Optical Microscopy-Guided Ion Beam Thinning for Precise Cryo Sample Targeting
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Solution Overview
Problem
Current ion beam thinning methods for cryo-electron tomography samples are 'blind cutting' and lack precision, as they cannot accurately identify or target specific structures within the sample, leading to inaccurate and complex sample preparation processes.
Innovation Solution
A method using optical microscopy imaging to mark a sample with an ion beam registration reference pattern, perform three-dimensional imaging, and project images to determine the position of the target for precise ion beam cutting, eliminating the need for fiducial markers and improving accuracy.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If ion beam thinning is performed without optical microscopy guidance, then the thinning process can be completed, but the positioning accuracy and ability to target specific structures is poor
Solution Approach 1:
The patent combines optical microscopy imaging system with ion beam processing system into an integrated platform. The optical microscope and ion beam column are merged in a single chamber, allowing simultaneous visualization and processing. This integration enables direct optical guidance of ion beam cutting without requiring separate alignment procedures or fiducial markers, thereby improving positioning accuracy while avoiding the complexity of multi-system coordination.
Solution Approach 2:
The patent introduces optical microscopy imaging as an intermediary between sample preparation and ion beam processing. The optical system provides real-time visualization of sample structures, acting as a mediator that guides the ion beam to specific targets. This intermediary enables precise positioning by allowing operators to visually identify and target specific cellular structures before and during the thinning process.
2Measurement precision
If fiducial markers are added to the sample for registration, then image alignment between optical and electron microscopy can be achieved, but the sample preparation complexity increases
Solution Approach 1:
The patent enables the sample to serve itself for registration purposes. By performing optical microscopy imaging directly on the sample in its native state within the same chamber, the system eliminates the need for external fiducial markers. The sample's own structural features are used for alignment and registration, simplifying sample preparation while maintaining high registration accuracy through direct visual correlation.
Solution Approach 2:
The patent extracts and removes the fiducial marker component from the system. Instead of adding external markers to the sample, the methodology uses the sample's intrinsic structural features visible under optical microscopy for registration. This extraction eliminates the need for marker addition, reduction, and coordination steps, thereby simplifying sample preparation while maintaining registration accuracy through direct visual correlation of sample features.
3Ease of operation
If traditional blind cutting method is used, then the thinning process is simple, but the ability to accurately locate and cut specific targets is lost
Solution Approach 1:
The patent implements real-time optical feedback during the ion beam thinning process. The optical microscopy system continuously images the sample surface, providing visual feedback to operators about the cutting progress and target location. This feedback loop allows operators to adjust the ion beam position and cutting parameters in real-time based on visual observation, maintaining both operational simplicity and high cutting precision for targeting specific cellular structures.
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 method enables high-precision, nano-scale targeting and cutting of specific structures within the sample, reducing sample preparation complexity and improving the repeatability of ion beam processing, allowing for accurate thinning suitable for transmission electron imaging and electron tomography.
Implementation Method 1
marking a surface of a sample using an ion beam, so as obtain a registration reference pattern
Implementation Method 2
performing a three-dimensional optical imaging on the sample to obtain a first three-dimensional light microscopy image
Implementation Method 3
performing an ion beam imaging on the sample to obtain an image excited by the ion beam
Data Source
AI summary
Provided is a method of ion beam processing based on optical microscopy imaging, including: marking a surface of a sample using the ion beam, so as obtain a registration reference pattern; performing a three-dimensional optical imaging on the sample to obtain a first three-dimensional light microscopy image; projecting the first three-dimensional light microscopy image to a cutting angle of the ion beam, and determining a first position of a to-be-researched target in the first three-dimensional light microscopy image based on the registration reference pattern; performing an ion beam imaging on the sample to obtain an image excited by the ion beam, and determining, according to the first position, a second position of the to-be-researched target in the image excited by the ion beam; and thinning the sample according to the second position to obtain a first slice containing the to-be-researched target.