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Real-time fluorescence monitoring frozen focused ion beam processing device and method

A technology of cryofocusing and real-time fluorescence, which is applied in measuring devices, fluorescence/phosphorescence, and material analysis through optical means. It can solve problems such as low processing efficiency, poor positioning accuracy, and low success rate, and improve processing accuracy. Processing process, the effect of improving processing efficiency and success rate

Pending Publication Date: 2021-11-19
INSITUTE OF BIOPHYSICS CHINESE ACADEMY OF SCIENCES
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0006] In view of the above-mentioned shortcomings and deficiencies of the prior art, the present invention provides a real-time fluorescence monitoring cryofocused ion beam processing device and method, which solves the problems of long processing time, low processing efficiency and poor positioning accuracy during ion beam processing of frozen samples in the prior art. , and the problem of low success rate

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  • Real-time fluorescence monitoring frozen focused ion beam processing device and method
  • Real-time fluorescence monitoring frozen focused ion beam processing device and method
  • Real-time fluorescence monitoring frozen focused ion beam processing device and method

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Embodiment 1

[0084] see Figure 1 to Figure 4 and Figure 11 , this embodiment provides a cryofocused ion beam processing device for real-time fluorescence monitoring, including a vacuum chamber 1 , an ion beam system 6 and a fluorescence imaging system 2 . Wherein, the vacuum chamber 1 is provided with a cold stage 3, and the cold stage 3 is used to store a frozen sample 8. A fluorescent marker is arranged in the frozen sample 8. When the excitation light is irradiated on the frozen sample 8, the fluorescent marker is excited. capable of emitting fluorescence. The ion beam system 6 is arranged on the vacuum chamber 1 and directed to the frozen sample 8 for emitting a focused ion beam to perform ion beam cutting on the frozen sample 8 . The fluorescence imaging system 2 is set on the vacuum chamber 1 and below the frozen sample 8, and is used for emitting excitation light to the frozen sample 8 and performing real-time optical imaging on the frozen sample 8 at the same time. In specific...

Embodiment 2

[0102] Based on Embodiment 1, compared with Embodiment 1, the real-time fluorescence monitoring cryofocused ion beam processing device provided in this embodiment differs only in the structure of the vacuum chamber hatch 120' and the structure of the cold stage 3 in this embodiment. different form of refrigeration.

[0103] see Figure 5 to Figure 10 , the vacuum chamber hatch 120' in this embodiment is a concave structure, and is provided with a sample transmission window 121'. The cold stage 3 is one end of the cryo-transfer device 7 , which includes a cryo-transfer rod 710 , a sample transfer tube 720 , a bellows assembly 730 , a cryo-transfer three-dimensional translation stage 740 and an angle adjustment device 760 . The sample transmission tube 720 is sealed and connected to the position of the sample transmission window 121 ′ through the bellows assembly 730 . One end of the frozen sample transmission rod 710 is a frozen sample fixing end 713 for fixing the frozen sam...

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Abstract

The invention relates to a real-time fluorescence monitoring frozen focused ion beam processing device and method. The real-time fluorescence monitoring frozen focused ion beam processing device comprises a vacuum chamber, an ion beam system and a fluorescence imaging system. A cold table is arranged in the vacuum chamber and is used for placing a frozen sample, and a fluorescent marker is arranged in the frozen sample. The ion beam system is arranged on the vacuum chamber, points to the frozen sample and is used for carrying out ion beam processing on the frozen sample. The fluorescence imaging system is arranged on the vacuum chamber, is positioned below the frozen sample and is used for carrying out real-time optical imaging on the frozen sample. During processing, exciting light is emitted to the frozen sample through the fluorescence imaging system, the fluorescence marker in the sample is excited to emit fluorescence, and the fluorescence is received by the fluorescence imaging system to form an image of a target structure in the frozen sample. By monitoring and analyzing fluorescence images and signals, the processing process of the frozen sample is monitored in real time, in-situ real-time monitoring and processing of the frozen sample are achieved, transfer is not needed, the processing accuracy of a target structure is improved, and the processing time is shortened.

Description

technical field [0001] The invention relates to the technical field of microscopic imaging, in particular to a real-time fluorescence monitoring cryo-focused ion beam processing device and method. Background technique [0002] Cryo-electron microscopy technology freezes biological samples in a near-physiological state through rapid freezing or high-pressure freezing technology to preserve high-resolution structures. Electron tomography is an important technical means of cryo-electron microscopy. Through a series of tilting imaging of frozen samples, three-dimensional reconstruction is performed to obtain a high-resolution three-dimensional structure of the ultrastructure of biological samples. Due to the transmission of samples, only tens to hundreds of nanometers thick samples can be observed. For cell tissue samples with a thickness of several microns or even thicker, it is impossible to observe directly and need to be sliced. There are two main types of techniques for ...

Claims

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Application Information

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IPC IPC(8): G01N23/223G01N23/04G01N21/64G01N21/01
CPCG01N23/223G01N23/04G01N21/6456G01N21/01G01N2021/6463G01N2021/0112
Inventor 季刚李硕果孙飞
Owner INSITUTE OF BIOPHYSICS CHINESE ACADEMY OF SCIENCES