Continuous Scanning Localization Microscopy Field of View Expansion
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Solution Overview
Problem
Conventional microscopy techniques face limitations in field of view (FOV) due to camera size, bandwidth, illumination intensity, and uniformity, leading to artifacts when stitching images together to achieve larger FOVs, which degrades image quality and increases memory and processing requirements.
Innovation Solution
A method of continuous scanning with a localization microscope, where the sample is repositioned to capture multiple image frames with modified FOV coordinates, allowing for the generation of composite images covering larger spatial regions without artifacts, using techniques like single-molecule switching (SMS) or single-molecule localization microscopy (SMLM), and accounting for motion-blur and sub-frame shifts.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Area of stationary object
If conventional techniques repeatedly image the sample then shift the FOV in discrete steps to achieve larger FOV, then the field of view is expanded, but artifacts are generated that degrade image quality
Solution Approach 1:
The patent implements continuous scanning of the sample through the field of view instead of discrete step-wise imaging. The sample is continuously translated while the camera captures images at a high frame rate, creating a continuous dataset that eliminates stitching artifacts between discrete frames. This continuous action maintains image quality while achieving expanded field of view.
Solution Approach 2:
The system dynamically adjusts the scanning parameters including translation speed, camera frame rate, and illumination intensity to optimize image quality across the expanded field of view. The continuous motion allows real-time adaptation of imaging parameters to maintain consistent image quality throughout the scanning process.
2Area of stationary object
If conventional techniques repeatedly image the sample then shift the FOV in discrete steps to achieve larger FOV, then the field of view is expanded, but memory storage and processing capabilities are exceeded
Solution Approach 1:
The continuous scanning data is segmented into manageable processing units based on temporal and spatial characteristics. The patent processes the continuous image stream in frames or frame groups, applying localization algorithms to each segment independently while maintaining continuity information. This segmentation reduces memory requirements compared to storing all discrete frames simultaneously.
Solution Approach 2:
The system performs preliminary processing of the continuous image stream by identifying and tracking localizable objects in real-time as images are captured. This preliminary action reduces the data volume that requires storage and post-processing, as object positions and trajectories are extracted during acquisition rather than from complete frame sets.
3Area of stationary object
If discrete step-wise FOV shifting is used to achieve larger FOV, then the field of view is expanded, but stitching artifacts are generated
Solution Approach 1:
The patent eliminates stitching operations entirely by using continuous scanning where the sample moves continuously through the field of view while the camera captures images at a high frame rate. The continuous nature of the scan creates an overlapping temporal and spatial dataset that can be assembled without discrete stitching, preventing the generation of stitching artifacts at frame boundaries.
Solution Approach 2:
The system performs preliminary registration and coordinate transformation during the continuous scanning process itself, mapping each captured frame to the global coordinate system based on the known continuous translation trajectory. This preliminary action ensures seamless integration of all frames without requiring post-acquisition stitching operations.
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 enables artifact-free imaging of large samples with improved quality and throughput by dynamically adjusting the FOV and focal position, overcoming limitations imposed by optical hardware and reducing processing demands.
Implementation Method 1
fluorescence or localization microscopy images
Data Source
AI summary
One aspect of the invention provides a method of continuously scanning with a localization microscope. The method includes: modifying a position of a sample relative to a field of view (FOV) of the localization microscope to capture a plurality of image frames of the sample, each captured image frame having a limited FOV; acquiring image frames with the localization microscope during at least one position modification; determining a set of localization position coordinates for at least one localizable object in the sample within at least one image frame of the plurality of image frames; determining one or more field of view (FOV) position coordinates for the at least one image frame; and modifying the set of localization position coordinates based on the one or more FOV position coordinates to produce a collection of coordinates covering a larger spatial region than the at least one image frame.