Preparation of cryogenic sample for charged-particle microscopy
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Solution Overview
Problem
Existing methods for preparing biological specimens in charged-particle microscopy often result in significant contamination due to rapid freezing, which hinders analysis by creating scattering sites for the imaging beam.
Innovation Solution
A method involving the simultaneous administration of cryogenic fluid to both sides of the sample through conduits, ensuring symmetric cooling and reducing contamination by preventing water leaching and ice crystal formation, while maintaining the sample in a static position to avoid damage to brittle grids or membranes.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Temperature
If rapid freezing is used to vitrify the sample, then the sample can be maintained at cryogenic temperature for microscopy, but significant contamination occurs due to water leaching and ice crystal formation
Solution Approach 1:
The sample is cooled from both sides simultaneously through separate conduits, dividing the cooling action into two symmetric segments that work together to achieve uniform vitrification without contamination
Solution Approach 2:
The cooling is applied locally at both surfaces of the sample through positioned conduits, ensuring that each region of the sample receives optimized cooling to prevent ice crystal formation and water leaching
2Temperature
If the sample is moved during cooling, then cooling can be applied, but brittle grids or membranes may be damaged
Solution Approach 1:
The sample holder itself serves as the cooling delivery system, with conduits integrated into the holder structure, allowing cooling to be applied without moving the fragile sample or its support grid
Solution Approach 2:
The conduits are pre-positioned and the cooling system is prepared before the sample is placed, allowing the sample to be cooled in its static, supported position without subsequent movement that could cause damage
3Temperature
If asymmetric cooling is applied to the sample, then cooling can be achieved, but uniform vitrification is not obtained
Solution Approach 1:
The invention deliberately uses symmetric rather than asymmetric cooling by applying cryogenic fluid to both sides of the sample through equally positioned conduits, ensuring uniform temperature distribution and vitrification throughout the sample
Solution Approach 2:
The cooling system is designed to create equipotential cooling conditions across the sample by positioning conduits symmetrically and delivering equal amounts of cryogenic fluid to both sides, eliminating temperature gradients that would cause non-uniform vitrification
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 significantly reduces contamination and achieves more uniform vitrification, allowing for clearer imaging and improved analysis in charged-particle microscopes by ensuring consistent and symmetric cooling of the sample.
Implementation Method 1
the sample is subjected to rapid cooling using a cryogen
Implementation Method 2
ensuring symmetric cooling and reducing contamination by preventing water leaching and ice crystal formation
Implementation Method 3
A cooling device, for maintaining the sample at a cryogenic temperature at least while it is on said sample holder
Data Source
Figure 1A
Figure 1B
Figure 1C
AI summary
A method of preparing a sample for study in a charged-particle microscope, whereby the sample is subjected to rapid cooling using a cryogen, comprising the following steps: - Providing two conduits for transporting cryogenic fluid, each of which conduits opens out into a mouthpiece, which mouthpieces are arranged to face each other across an intervening gap; - Placing the sample in said gap; - Pumping cryogenic fluid through said conduits so as to concurrently flush from said mouthpieces, thereby suddenly immersing the sample in cryogenic fluid from two opposite sides.