Non-Coaxial Scanning Microscope Multiplexing
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Solution Overview
Problem
Current scanning microscopy techniques, such as confocal laser scanning microscopy, face limitations in multiplexing capability due to coaxial illumination designs, which restrict the number of fluorescent agents that can be detected simultaneously and lead to crosstalk issues, reducing throughput and scalability.
Innovation Solution
The implementation of a scanning optical microscope with non-coaxial illumination light beams from multiple sources, allowing each beam to propagate at different angles, enabling simultaneous scanning and detection of fluorescent emissions at various wavelengths, decoupling focal spots for independent detection paths and modular expansion.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Device complexity
If coaxial illumination design is used, then device complexity is reduced, but multiplexing capability is limited and crosstalk occurs
Solution Approach 1:
The illumination system is segmented into multiple independent non-coaxial beam paths, each capable of delivering light at a specific wavelength to a distinct focal location. This segmentation allows simultaneous illumination at multiple wavelengths without interference, resolving the contradiction between device simplicity and multiplexing capability.
Solution Approach 2:
The patent transitions from a single-axis (coaxial) illumination geometry to a multi-dimensional non-coaxial arrangement where beams propagate at different angles. This dimensional change enables spatial separation of focal spots while maintaining a relatively simple optical architecture, thereby improving multiplexing capability without proportionally increasing device complexity.
2Adaptability or versatility
If multiple laser sources are added to increase multiplexing capability, then fluorescent detection capability improves, but device complexity and crosstalk increase
Solution Approach 1:
Each laser source is assigned a dedicated non-coaxial beam path with its own focal spot, creating independent detection channels. This segmentation prevents crosstalk between channels while allowing simultaneous operation of multiple lasers, thereby enhancing fluorescent detection capability without proportionally increasing device complexity.
Solution Approach 2:
The non-coaxial beam geometry acts as an intermediary mechanism that couples each laser source to its specific focal region while naturally isolating it from other beams. This intermediary arrangement enables multiple laser sources to operate simultaneously without requiring complex isolation optics or sequential operation, thus improving detection capability while controlling complexity.
3Adaptability or versatility
If non-coaxial illumination beams are used, then multiplexing capability and scalability improve, but device complexity increases
Solution Approach 1:
The optical system is divided into modular non-coaxial beam modules, each handling a specific wavelength channel. This modular segmentation allows the system to scale by simply adding or removing modules without redesigning the entire optical path, thereby improving multiplexing capability and scalability while keeping the complexity of each individual module manageable.
Solution Approach 2:
The non-coaxial beam architecture provides a universal platform that can accommodate multiple laser sources and detection configurations through a single integrated optical system. This multi-functional design allows the same basic structure to support various multiplexing schemes, improving versatility without requiring separate complex systems for each function.
4Ease of operation
If coaxial illumination is used, then optical path alignment is simplified, but throughput and detection efficiency decrease
Solution Approach 1:
The optical detection system is segmented into multiple independent detection paths, each aligned with a specific non-coaxial illumination beam and its corresponding focal spot. This segmentation allows each detection channel to be optimized independently for maximum throughput while maintaining relatively simple alignment procedures for each module, thereby resolving the contradiction between alignment simplicity and detection efficiency.
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 enhances scalability, maintains constant throughput regardless of the number of laser sources, and allows for a broader selection of fluorescent agents, improving the multiplexing capability and simplifying the device design.
Implementation Method 1
Each probe laser beam at a respective laser wavelength excites generation in the sample of a corresponding fluorescent emission at different fluorescent emission wavelengths caused by laser light at the respective laser wavelength
Implementation Method 2
an optical beam routing device located in the common optical path of the combined probe laser beam bundle having the different probe laser beams and structured to include an optical input port to receive the combined probe laser beam bundle to redirect the combined probe laser beam bundle and a different optical output port
Implementation Method 3
an optical scanning device located to receive the combined probe laser beam bundle having the different probe laser beams and structured to include an actuator to cause a scanning of the combined probe laser beam bundle having the different probe laser beams to output the different probe laser beams with each being scanned over a scanning range while still being in different beam propagation angles relative to one another
Implementation Method 4
a microscope objective lens placed in an optical path of the combined probe laser beam bundle having the different probe laser beams between the optical scanning device and the sample stage to focus the different probe laser beams onto the sample surface at different probe laser beam locations near or on the sample surface
Implementation Method 5
each optical detection module includes an optical diffractive device to spatially separates different optical spectral components of a received fluorescent emission beam into different fluorescent emission spectral beams at different optical fluorescent emission wavelengths
Data Source
AI summary
A scanning microscope multiplexes illumination light beams at different optical wavelengths (e.g., from different light sources) by placing illumination light beams non-coaxially to create separate focuses at specimen. Reflected or fluorescent light generated by the different illumination light beams is recorded simultaneously while scanning the specimen.