Inverted Microscope Merging TIRF and Confocal Systems
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Solution Overview
Problem
Current microscope systems, such as total internal reflection fluorescence microscopy and disk scanning confocal microscopy, have limitations in observing the propagation of excitatory states in cells, particularly due to restricted observation ranges and inability to detect variations in excitation states beyond the reach of evanescent waves or confocal sections.
Innovation Solution
An inverted microscope system combining a total internal reflection fluorescence microscopy optical system and a disk scanning confocal optical system, with a rotary disk and confocal openings, allows for adjustable relative distances between the objective lens and the confocal position, enabling extended observation and sectioning of fluorescence across multiple layers.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If a total internal reflection fluorescence microscopy optical system is used to illuminate only the specimen near the glass interface, then the observation depth is limited to several hundred nm, but the ability to observe excitatory state propagation in upper layer cells is improved
Solution Approach 1:
The patent combines the total internal reflection fluorescence microscopy optical system and the disk scanning confocal optical system into a single integrated system. The total internal reflection system provides high-precision observation at the glass interface, while the confocal system extends observation capability to upper layer cells, resolving the contradiction between observation depth precision and observation range.
Solution Approach 2:
The integrated optical system serves multiple functions: it can perform total internal reflection fluorescence microscopy for high-resolution interface observation, disk scanning confocal microscopy for extended depth observation, and simultaneously detect excitatory state propagation across multiple cell layers, achieving versatility while maintaining precision.
2Measurement precision
If a disk scanning confocal optical system is used to observe fluorescence from specimen surface in real time, then the sectioning capability is improved, but the ability to detect intensity variations across multiple layers is limited
Solution Approach 1:
By merging the disk scanning confocal system with the total internal reflection system, the patent achieves both real-time sectioning capability and multi-layer detection capability. The confocal system provides optical sectioning while the integrated system detects intensity variations across multiple cell layers through coordinated observation.
Solution Approach 2:
The patent extends observation from a single focal plane to multiple layers by utilizing the vertical dimension. The adjustable focus position allows the system to scan through different depths and detect intensity variations across multiple cell layers, adding a depth dimension to the observation capability.
3Device complexity
If the focus position and confocal position are fixed relative to each other, then the system structure is simplified, but the ability to adjust observation depth and detect variations across layers is reduced
Solution Approach 1:
The patent introduces dynamic adjustability to the system by allowing the focus position of the objective lens to be changed relative to the confocal position of the rotary disk. This dynamic adjustment capability enables flexible depth control and multi-layer observation while maintaining a relatively simple overall system structure.
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 combination enhances the observation range and sectioning capability, allowing for real-time detection of excitatory state propagation and intensity variations across multiple layers, improving the understanding of cellular excitatory states and reducing photobleaching.
Implementation Method 1
an objective lens (31) that collects at least observation light from the specimen S
Implementation Method 2
a tube lens (32) that forms an image using the observation light collected by the objective lens (31)
Implementation Method 3
illumination light is totally reflected from a glass interface on which a specimen is placed to illuminate only the specimen just near the glass interface using an evanescent wave generated by the total reflection illumination
Implementation Method 4
illuminate only the specimen just near the glass interface using an evanescent wave generated by the total reflection illumination, and thus the generated fluorescence can be observed
Implementation Method 5
the disk scanning confocal optical system is capable of observing only the fluorescence from the specimen surface in real time by spinning, at a high speed, a disk on which a confocal opening such as a slit or a pinhole is placed
Data Source
AI summary
An inverted microscope system includes an objective lens holding unit that holds an objective lens configured to collect at least observation light from a specimen, a tube lens configured to form an image using the observation light collected by the objective lens, a total internal reflection fluorescence microscopy optical system provided between the objective lens and the tube lens and configured to observe the observation light from the specimen using a total reflection illumination, and a disk scanning confocal optical system including a rotary disk on which a confocal opening is formed, the confocal opening being placed at a position substantially conjugate to a focus position of the objective lens. A relative distance between the focus position of the objective lens and the substantially conjugate position is changeable along an optical path of the observation light.