Segmented Beam Splitter for Multi-Spot Microscopy
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Solution Overview
Problem
Existing laser scanning microscopes struggle to simultaneously excite multiple fluorescent proteins with overlapping excitation spectra without significant signal loss due to beam splitter limitations, and they require reconfiguration for multicolor imaging, which is time-consuming.
Innovation Solution
A device with a segmented beam splitter that is laterally subdivided into multiple filter fields allows for the selection of wavelength ranges and/or polarization directions for partial illumination and detection beams, enabling simultaneous excitation of multiple fluorescent proteins without signal loss and eliminating the need for reconfiguration.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Use of energy by moving object
If a beam splitter with high reflectivity at 594 nm is used to excite mCherry, then the excitation efficiency of mCherry is improved, but significant parts of the emission from tdTomato are blocked
Solution Approach 1:
The beam splitter is segmented into multiple filter fields, each optimized for specific wavelength ranges. This allows different regions of the beam splitter to handle different spectral channels simultaneously, enabling efficient excitation of multiple fluorophores without cross-blocking of emission signals.
Solution Approach 2:
Different regions (filter fields) of the beam splitter are assigned different optical properties tailored to specific wavelength ranges. Each filter field has optimized reflectivity and transmission characteristics for its designated spectral range, allowing local optimization of excitation efficiency while preserving emission signals from other channels.
2Use of energy by moving object
If a beam splitter with high reflectivity at 561 nm is selected for tdTomato excitation, then the excitation of tdTomato is improved, but the device must be reconfigured for multicolor imaging, which takes several seconds
Solution Approach 1:
The beam splitter is divided into multiple filter fields that can be independently activated. This segmentation allows the system to switch between different excitation configurations by activating different filter fields without physical reconfiguration of the entire beam splitter, reducing reconfiguration time from several seconds to minimal time.
Solution Approach 2:
The beam splitter is designed with dynamic switching capability between different filter fields. This allows the optical path to be dynamically reconfigured by changing which filter fields are active, enabling rapid switching between different excitation wavelengths for multicolor imaging without mechanical repositioning.
3Measurement precision
If the spectral bandwidths of beam splitters are kept narrow (10-30 nm) to achieve spectral discrimination, then the spectral resolution is improved, but the transmission window to fluorescence emission is blocked
Solution Approach 1:
The beam splitter is segmented into multiple filter fields, each with optimized spectral characteristics. This segmentation allows each filter field to have narrow bandwidth for spectral discrimination in its specific range, while collectively providing adequate transmission windows for fluorescence emission across all measured wavelength ranges.
Solution Approach 2:
Each filter field is designed with local spectral optimization, having narrow bandwidth and high reflectivity for its specific excitation wavelength range while maintaining transmission windows for corresponding fluorescence emission. This local quality approach resolves the contradiction by optimizing each region for its specific function rather than using a uniform design.
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 solution enhances the spectral excitation performance of laser scanning microscopes, allowing for simultaneous and efficient excitation of multiple pigments, particularly in the red wavelength range, while achieving simultaneous data recording without the need for reconfiguring the device.
Implementation Method 1
at least one beam splitter which is segmented laterally to a surface normal of the beam splitter into at least two filter fields for the selection of wavelength ranges and/or polarization directions
Data Source
AI summary
A device and a method for multi-spot scanning microscopy using a segmented color separator are disclosed, wherein a beam splitter is segmented laterally to a surface normal of the beam splitter into at least two filter fields for the selection of wavelength ranges and/or polarization directions. Individual partial illumination beams are thus each guided in an illumination beam path into a light spot on or in a simple to be examined and scanned over it. Detection light, which the sample emits in partial detection means after irradiation using the individual partial illumination beams, is guided onto a detection unit and detected thereby, wherein the partial illumination beams and/or the partial detection beams are each selected according to wavelength ranges and/or polarization directions by means of the beam splitter segmented laterally to a surface normal.