Digital Parallel Frequency Fluorometry for Fast Lifetime Imaging
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Solution Overview
Problem
Existing fluorescence lifetime imaging microscopy (FLIM) devices are cumbersome to integrate with fluorescence intensity imaging systems and are limited by high cost, low portability, and inefficiency in resolving picosecond lifetimes, especially in miniaturized microscopy applications where temporal resolution is restricted by sample brightness rather than instrument capabilities.
Innovation Solution
A digital parallel acquisition method for multifrequency phase fluorometry that eliminates the need for modulating detector gain, allowing for simultaneous acquisition of multiple frequencies with 100% duty cycle, reducing data acquisition time, and enabling the use of various light detectors, including PMTs, MCPs, and APDs, without hardware modifications or significant power consumption.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If traditional frequency domain fluorometry is used to measure fluorescence decay times, then measurement precision can be achieved, but data acquisition time is excessively long (minutes to hours)
Solution Approach 1:
The patent divides the measurement process into multiple frequency channels (e.g., 16 parallel channels) that simultaneously measure different harmonic frequencies of the excitation light. This segmentation allows the system to extract multiple decay time components in parallel rather than sequentially, reducing acquisition time from minutes to seconds while maintaining measurement precision through multi-frequency analysis
Solution Approach 2:
The patent employs periodic modulation of the excitation light source at multiple frequencies to encode fluorescence decay information across different time scales. By using periodic action at harmonically related frequencies, the system can resolve multiple exponential decay components simultaneously, achieving both high precision and fast acquisition through frequency-domain multiplexing
2Measurement precision
If detector gain is modulated in traditional frequency domain instruments, then phase and modulation measurements can be obtained, but device complexity and power consumption increase
Solution Approach 1:
The patent replaces the mechanical/electronic detector gain modulation system with a digital signal processing approach. Instead of physically modulating the detector gain using complex hardware circuits, the system uses software-based digital demodulation of the photodetector output signal, significantly reducing device complexity and power consumption while preserving the ability to measure phase and modulation accurately
Solution Approach 2:
The patent introduces digital signal processing algorithms as an intermediary between the photodetector and the measurement output. These algorithms perform digital demodulation and phase extraction, acting as a virtual mediator that replaces physical detector modulation hardware and enables precise measurements through computational methods rather than complex electronic modulation circuits
3Measurement precision
If multiple frequencies are measured sequentially in traditional instruments, then accurate decay time determination can be achieved, but productivity is severely limited
Solution Approach 1:
The patent merges multiple frequency measurements into a single simultaneous acquisition by using parallel detection channels that process different harmonic frequencies concurrently. This merging of measurement operations into a unified parallel system maintains the accuracy of individual frequency measurements while achieving high productivity through simultaneous multi-frequency analysis
Solution Approach 2:
The patent creates a universal measurement system that can handle multiple decay time components across different frequency ranges using the same hardware platform. The parallel channel architecture provides multi-functionality, allowing the instrument to measure various fluorescence lifetimes simultaneously without requiring separate measurement sequences, thereby enhancing both precision and productivity
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 significantly reduces data acquisition time from minutes to seconds, increases measurement accuracy, and allows for portable, cost-effective implementation in biomedical and clinical applications, while maintaining high sensitivity and precision in determining fluorescence decay times.
Implementation Method 1
Fluorescence is the light emitted by molecules in solution (or in a solid or gaseous state) following the absorption of radiation
Implementation Method 2
The emission light 102 is phase-shifted and demodulated with respect to the excitation light 104
Implementation Method 3
a photodetector to generate an output signal corresponding to the frequency and phase of the modulated fluorescence
Data Source
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AI summary
A system and method is provided for improved fluorescence decay time measurement. A digital heterodyning technique is disclosed in which a photon detector is sampled at a rate slightly faster than a digitally pulsed excitation signal. A resulting cross correlation frequency is low enough to be read by inexpensive electronics such as by a field programmable gate array. Phase information in the signal provides correlation with corresponding photon detections.