Tunable Wavelength Excitation Module for Multiphoton Microscopy
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Solution Overview
Problem
Conventional temporal focusing-based multiphoton excitation fluorescence microscopy systems face difficulties in adjusting the excitation wavelength in real time, limiting their ability to efficiently excite fluorophores with different two-photon absorption spectra, especially in multifluorophore imaging, which restricts the selection of fluorophores for biomedical applications and affects image quality.
Innovation Solution
A temporal focusing-based multiphoton excitation fluorescence microscopy system with a tunable-wavelength excitation module, comprising an angular adjustment unit, a control unit, and a diffraction unit, allows for real-time adjustment of the excitation wavelength by using a reflection device and optical grating to alter the output angle of the excitation light, enabling dynamic selection of suitable wavelengths for optimal imaging.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Adaptability or versatility
If a fixed-wavelength pulse laser is used for excitation, then the system structure is simple, but the system cannot efficiently excite fluorophores with different two-photon absorption spectra
Solution Approach 1:
The patent applies the dynamics principle by making the excitation wavelength tunable through a rotating angular adjustment unit. The reflection device can rotate to different angles, dynamically changing the wavelength of excitation light that reaches the diffraction unit and subsequently the sample. This allows the system to adapt to different fluorophore absorption spectra while maintaining a relatively compact structure.
Solution Approach 2:
The patent implements parameter changes by varying the wavelength of the excitation laser through mechanical adjustment. The angular adjustment unit changes the angle at which light strikes the diffraction unit, thereby selecting different wavelengths from the broadband laser source. This enables the system to match the two-photon absorption spectra of various fluorophores.
2Productivity
If the incident wavelength is adjusted in real time, then the multiphoton excitation efficiency is optimized, but the diffraction device separates frequencies at different diffraction angles making adjustment difficult
Solution Approach 1:
The patent introduces an angular adjustment unit as an intermediary between the fixed diffraction device and the sample. This intermediate component (the rotating reflection device) simplifies the operation by providing a straightforward rotational mechanism to select wavelengths, rather than requiring direct manipulation of the diffraction device itself. The control unit further mediates this process by automatically positioning the angular adjustment unit to the correct angle.
Solution Approach 2:
The patent replaces complex mechanical wavelength adjustment mechanisms with a simpler rotational angular adjustment unit. Instead of moving the diffraction grating itself or using complex optical path modifications, the system uses a single rotational degree of freedom to control which wavelength is directed to the sample, significantly easing operation.
3Adaptability or versatility
If a single excitation wavelength is used, then the system configuration is simple, but the selection of fluorophores for biomedical applications is restricted
Solution Approach 1:
The patent implements universality by designing a wavelength selection module that can accommodate multiple fluorophores with different absorption spectra using a single laser source. The angular adjustment unit combined with the diffraction unit creates a multi-functional excitation system that can be tuned to match various two-photon absorption spectra, expanding fluorophore selection without requiring multiple separate laser systems.
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 enables real-time adjustment of the excitation wavelength, resulting in higher image quality by effectively exciting multiple fluorophores with different absorption spectra, overcoming the limitations of fixed-wavelength systems and improving imaging depth in three-dimensional biospecimens.
Implementation Method 1
a diffraction unit for receiving the reflected excitation light and generating detecting excitation light
Implementation Method 2
an angular adjustment unit which is a reflection device configured to rotate at a fixed point in order to reflect the excitation light
Implementation Method 3
Multiphoton excitation (MPE) fluorescence microscopes can be used in the biomedical field
Data Source
AI summary
The invention provides a temporal focusing-based multiphoton excitation fluorescence microscopy system capable of tunable-wavelength excitation and an excitation wavelength selection module thereof. The temporal focusing-based multiphoton excitation fluorescence microscopy system comprises: an excitation light generating module for generating excitation light; an excitation wavelength selection module for generating reflected light having a predetermined output angle in accordance with the wavelength of the excitation light and generating detecting excitation light through a diffraction unit; and a fluorescent microscope. With the practice of the present invention, the wavelength of the excitation light can be changed by the excitation wavelength selection module in real time to excite different fluorophores with the corresponding maximum multiphoton absorption wavelengths, so as to obtain better fluorescent signals and improve the image quality of the temporal focusing-based multiphoton excitation fluorescence microscope system.