Sample Exchange Chamber for Air-Free Immunostaining Transfer
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Solution Overview
Problem
Immunoelectron microscopy is not widely adopted due to limitations in reagents for optical microscope sample preparation and immunostaining, and pre-fixatives and epoxy resins used in electron microscopy reduce antigenicity and complicate the staining process, making it difficult to observe osmium black and microscopic gold particles with scanning electron microscopes.
Innovation Solution
A sample exchange chamber with integrated staining, cleaning, evacuation, and sterilization mechanisms, along with an autoloader mechanism, allows for automated sample handling and immunostaining in a charged particle beam apparatus, enabling stable and simpler pathological diagnosis.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Stability of the object's composition
If pre-fixatives and post-fixatives are used to improve microstructure retention in electron microscopy, then the microstructure form is better preserved, but antigenicity is reduced and immune reaction is inhibited
Solution Approach 1:
The patent divides the sample preparation process into separate stages: optical microscope sample preparation (with immunostaining) is performed first, then the sample is transferred to electron microscope observation. This segmentation allows different fixation and staining protocols to be optimized for each microscopy type, avoiding the need to compromise antigenicity for electron microscopy fixation.
Solution Approach 2:
The patent performs immunostaining and DAB development as preliminary actions before electron microscopy observation. The DAB reaction product is formed first, then the sample undergoes osmium staining and other electron microscopy-specific treatments. This preliminary action ensures that the immune reaction occurs when antigenicity is still intact, before any electron microscopy fixatives are applied.
2Reliability
If conventional immunoelectron microscopy methods are used to improve staining stability, then the staining becomes more stable, but the procedure becomes complicated and observation of osmium black and gold particles becomes difficult with SEM
Solution Approach 1:
The patent employs an automated sample transfer system that automatically transfers samples between optical and electron microscopes without manual intervention. The system self-manages the complex multi-step process of immunostaining, DAB development, osmium staining, and sample transfer, reducing procedural complexity and human error while maintaining staining stability.
Solution Approach 2:
The patent combines multiple functions into an integrated system that performs both optical and electron microscopy observations on the same sample platform. The sample stage and transfer mechanisms are merged into a single automated system, allowing seamless transition between microscopy types without separate preparation procedures.
3Measurement precision
If TEM is used to achieve high resolution for immunoelectron microscopy, then observation resolution is improved, but facility cost and space requirements increase significantly
Solution Approach 1:
The patent develops a sample preparation and observation system that is universal for both optical and electron microscopy. The same sample stage, transfer mechanism, and chamber system serve both microscopy types, allowing institutions to perform high-resolution immunoelectron microscopy using more accessible SEM facilities rather than requiring dedicated TEM infrastructure.
4Ease of operation
If samples are exposed to air during preparation and observation, then sample accessibility is improved, but sample contamination and degradation occur
Solution Approach 1:
The patent uses a sealed chamber system that maintains an inert or controlled atmosphere during sample preparation and transfer. The chamber is evacuated or filled with protective gas, preventing sample contamination from air exposure while allowing automated manipulation and observation. This inert environment protects samples throughout the multi-step preparation and transfer process.
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
Facilitates automated staining, observation, and analysis in a scanning electron microscope, reducing labor and time for sample preparation and providing clear contrast for accurate identification of positive sites without exposing samples to air.
Implementation Method 1
an immunostaining method, which is one of the staining methods, is a procedure for identifying functions of the tissue and organelles by visualizing expression of specific genes and various marker proteins mainly using an antigen-antibody reaction which is a specific reaction between an antigen substance such as protein and an antibody
Implementation Method 2
a cleaning mechanism configured to clean the sample
Implementation Method 3
an evacuation mechanism configured to evacuate the container
Implementation Method 4
a sterilization mechanism configured to sterilize the sample and inside of the container
Implementation Method 5
a charged particle beam apparatus includes: a sample chamber; a sample exchange chamber... and an autoloader mechanism configured to transfer the sample between the sample exchange chamber and the sample chamber
Data Source
AI summary
In order to enable use of novel immunostaining for a pathological diagnosis and research, the invention provides a sample exchange chamber including: a container 2 into which a substrate on which a sample is placed can be introduced; a specific solution inlet 3, which is a staining mechanism that stains the sample; a cleaning liquid inlet 4, which is a cleaning mechanism that cleans the sample; an evacuation port 5, which is an evacuation mechanism that evacuates the container; a drain port 6; and a sterilization mechanism 7 that sterilizes the sample and inside of the container.