Multiplexed One-Photon and Nonlinear Microscopy Alignment
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Solution Overview
Problem
Current microscopy techniques, such as one-photon and two-photon fluorescence microscopy, have limitations in terms of cross-modality alignment, resolution, and depth penetration, making it challenging to align and compare images from different modalities, especially for biological tissues.
Innovation Solution
The development of a multiplexing and demultiplexing system that combines one-photon and nonlinear microscopy, enabling fast temporal multiplexing and spatial alignment of tissue images, thereby overcoming the limitations of individual modalities.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Area of stationary object
If one-photon fluorescence microscopy is used, then a large field of view and scalability are achieved, but resolution, contrast, and depth of tissue penetration are limited
Solution Approach 1:
The patent combines one-photon and two-photon microscopy systems into a single integrated platform, allowing both modalities to operate simultaneously or sequentially. This merging enables the system to achieve both large field of view (from one-photon) and high resolution (from two-photon) without requiring separate apparatuses, directly resolving the contradiction between field of view and resolution.
Solution Approach 2:
The microscopy system is designed with multi-functionality, incorporating both one-photon and two-photon imaging capabilities, as well as alignment features, within a single device. This universal platform can adapt to different imaging requirements - providing wide-field views when needed and high-resolution deep-tissue imaging when required, thereby eliminating the trade-off between field of view and resolution.
2Measurement precision
If two-photon fluorescence microscopy is used, then resolution and depth of tissue penetration are improved, but field of view is limited and the apparatus is large
Solution Approach 1:
By merging one-photon and two-photon microscopy capabilities into a single system, the patent enables the user to switch between wide-field one-photon imaging and high-resolution two-photon imaging based on the specific application requirements. This integration allows the system to provide both large field of view and high resolution without compromising either capability.
Solution Approach 2:
The system incorporates dynamic switching capabilities between one-photon and two-photon modes, allowing real-time adaptation to different imaging needs. The system can dynamically adjust its operating mode to provide either wide-field coverage or high-resolution imaging depending on the sample and experimental requirements, thereby resolving the fixed trade-off between field of view and resolution.
3Ease of operation
If one-photon imaging is used, then portability is achieved, but cross-modality alignment with two-photon imaging becomes impossible
Solution Approach 1:
The patent merges alignment capabilities directly into the portable one-photon microscopy system, enabling the same device to perform both imaging and alignment functions. By integrating the alignment mechanism within the portable system, the patent eliminates the need for separate alignment apparatuses and maintains portability while enabling cross-modality alignment with two-photon imaging data.
Solution Approach 2:
The system uses fluorescent beads as intermediary markers that can be visualized in both one-photon and two-photon modalities. These beads serve as a common reference framework that mediates between the two imaging techniques, enabling accurate registration and alignment of images from different modalities while maintaining the portability of the one-photon system.
4Adaptability or versatility
If separate one-photon and two-photon microscopy systems are used, then each modality can be optimized independently, but cross-modality alignment and comparison become challenging
Solution Approach 1:
The patent combines multiple microscopy modalities and alignment functions into a single integrated system, reducing the overall complexity compared to operating separate systems. By merging the functionalities, the system eliminates the need for multiple separate apparatuses while maintaining the optimization benefits of each modality, thereby reducing device complexity and enabling seamless cross-modality alignment.
Solution Approach 2:
The integrated system provides multi-functionality, accommodating both one-photon and two-photon imaging modes along with alignment capabilities in a single platform. This universal design allows the system to perform all required functions - imaging in different modalities and aligning them - without requiring separate specialized systems, thereby reducing overall system complexity while maintaining adaptability.
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 allows for precise alignment and comparison of images from different microscopy modalities, providing high-resolution, three-dimensional imaging capabilities while maintaining the portability and large field of view of one-photon microscopy.
Implementation Method 1
one-photon fluorescence microscopy... utilizes an incoherent source of light... to illuminate and collect light from the sample
Implementation Method 2
nonlinear optical effects such as two-, three-photon absorption... A light generated in such a process has a shorter wavelength than the excitation light
Implementation Method 3
cross-modality alignment becomes a serious challenge... finding the same fragment of tissue such as particular cells of interest becomes impossible
Data Source
AI summary
We combine linear and nonlinear microscopy modalities. This approach is capable of imaging a sample using the two modalities concurrently, generating time sequences of images of the sample having different properties characteristic of each of the modalities. We can separate light emitted from the sample, generated by either modality, even if the spectrum of light is identical for each modality. The nonlinear microscopy modality allows one to create high resolution volumetric images of the sample. The concurrent imaging of the sample using linear and nonlinear microscopy modalities enables one to correlate a spatial structure and temporal dynamics of the sample acquired in each modality allowing one to create a mapping between images across the modalities. The created mapping can be used to match images of a tissue sample acquired using linear modality in a living organism with high resolution, two- or three-dimensional images obtained using nonlinear microscopy modality.