Cryo Microscope Thermal Isolation via Coupling Medium
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Solution Overview
Problem
Current super-resolution cryogenic microscopy techniques face challenges in maintaining a temperature differential over small distances, leading to reduced imaging resolution and performance when using conventional light microscopes, as heat transfer between the sample and the microscope objective compromises the temperature difference.
Innovation Solution
A cooling microscope assembly with a first circulation system for the objective lens and a second circulation system for the sample stage, utilizing a coupling medium with low thermal conductivity to maintain a temperature differential between the objective lens and the sample, allowing for high-resolution imaging without compromising the microscope's performance.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If the sample is cooled to cryogenic temperatures to reduce molecular motion and bleaching rate, then imaging resolution is improved, but heat transfer to the objective lens compromises the temperature differential
Solution Approach 1:
A thermal intermediary layer with low thermal conductivity is introduced between the cryogenic sample stage and the room-temperature objective lens. This intermediary layer acts as a thermal barrier that prevents heat transfer from the warm objective to the cold sample, maintaining the temperature differential necessary for cryogenic imaging while allowing optical signals to pass through.
Solution Approach 2:
The microscope system is segmented into distinct thermal zones: a cryogenic sample stage maintained at low temperatures for imaging, and a room-temperature objective lens maintained at higher temperatures for optical performance. The segmentation allows each component to operate at its optimal temperature independently, with thermal isolation preventing unwanted heat transfer between zones.
2Temperature
If air gaps are used to insulate the objective lens from the cold sample, then thermal isolation is improved, but numerical aperture is reduced due to total internal reflection
Solution Approach 1:
An optical intermediary layer with refractive index matching the immersion fluid is introduced between the objective lens and the cryogenic stage. This intermediary layer serves dual functions: it maintains thermal isolation by having low thermal conductivity while providing optimal optical coupling by matching refractive indices, thereby preventing total internal reflection and maximizing numerical aperture.
Solution Approach 2:
The refractive index parameter of the intermediary layer is specifically optimized to match the immersion fluid (approximately 1.33-1.4), while its thermal conductivity is minimized to maintain thermal isolation. This parameter optimization allows the system to achieve both good thermal isolation and high numerical aperture simultaneously.
3Measurement precision
If the working distance is reduced to use higher NA lenses, then imaging resolution is improved, but maintaining temperature differential becomes more difficult
Solution Approach 1:
A thin film intermediary layer with low thermal conductivity is placed between the objective lens and the cryogenic stage. This thin film provides sufficient thermal isolation to maintain the temperature differential even at reduced working distances, while allowing the high NA lens to achieve its optimal performance without excessive heat transfer compromising the cryogenic environment.
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
The assembly achieves a significant increase in imaging resolution from 20 nm to 2 nm, enabling molecular-level resolution of protein molecules while maintaining a stable temperature differential, thus overcoming the limitations of existing cryogenic microscopy setups.
Implementation Method 1
utilizing a coupling medium with low thermal conductivity to maintain a temperature differential between the objective lens and the sample
Implementation Method 2
a second circulation system for a sample stage, comprising: an insulated enclosure, comprising: a cold block for sample placement and one or more ports for transport of a cooling fluid
Implementation Method 3
a first circulation system for an objective lens, comprising: a light transparent element and a collar, including an upper surface capable of sealing engagement with the objective lens and a lower surface capable of sealing engagement with the light transparent element, the upper surface of the collar and the light transparent element defining a region for housing immersion fluid
Data Source
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AI summary
New systems and methods are described for maintaining a desired steady state temperature differential between two objects that may otherwise undergo heat transfer to restore thermal steady state. In one application, a cooling microscope assembly and its use with conventional optical microscopes are described for achieving super-resolution imaging. The assembly allows for the high resolution imaging of samples at cryogenic temperatures while maintaining the temperature of the objective lens above freezing by employing circulation systems and a coupling fluid between the sample and objective Jens.