Fluorescence Lifetime Measurement via Temporal Averaging
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Solution Overview
Problem
Conventional methods for measuring fluorescence lifetime, such as time-correlated single photon counting (TCSPC), are limited by long measurement times, making it difficult to achieve high-speed imaging in fluorescence lifetime imaging microscopy (FLIM), especially for three-dimensional scanning microscopy applications.
Innovation Solution
An apparatus that generates pulsed excitation light and uses a light sensor to convert fluorescence photons into electrical signals, with a signal processor calculating the time average of these signals to determine the fluorescence lifetime, allowing for high-speed measurement by subtracting the apparatus delay time from the average time of the fluorescence signal.
Engineering Contradictions & Design Principles
Engineering Contradiction Analysis
1Measurement precision
If time-correlated single photon counting (TCSPC) is used to measure fluorescence lifetime, then measurement precision is improved, but measurement time increases significantly
Solution Approach 1:
The patent replaces the conventional TCSPC method with a direct temporal averaging method that calculates fluorescence lifetime by measuring the average time of detected photons and subtracting the instrument response function average time. This substitution eliminates the need for complex histogram accumulation and iterative fitting procedures, dramatically reducing measurement time while maintaining precision through the relationship: τ_fluorescence = <t_photon> - <t_IRF>
Solution Approach 2:
The patent changes the measurement approach from accumulating photon arrival times in histograms (TCSPC) to directly calculating temporal averages of detected signals. By focusing on the first moment of the fluorescence decay curve rather than full histogram reconstruction, the method achieves faster acquisition suitable for imaging applications while preserving measurement accuracy
2Measurement precision
If conventional fluorescence lifetime measurement methods are used, then measurement accuracy is maintained, but imaging speed decreases
Solution Approach 1:
The patent extracts only the essential temporal information needed for fluorescence lifetime measurement by calculating the average photon arrival time and subtracting the instrument response function average time. This extraction approach eliminates unnecessary data processing steps such as histogram binning and curve fitting, enabling rapid acquisition for imaging applications while maintaining measurement accuracy
Solution Approach 2:
The patent uses the instrument response function (IRF) measured from a reference sample to correct the fluorescence signal. By copying and subtracting the IRF average time from the measured photon average time, the method accurately removes instrumental broadening effects without requiring full deconvolution, thus maintaining precision while improving speed
3Measurement precision
If detailed spectroscopic information is collected for fluorescence lifetime imaging, then measurement precision is improved, but measurement time increases
Solution Approach 1:
The patent applies partial action by measuring only the first moment (average time) of the fluorescence decay curve rather than reconstructing the complete decay profile. This partial measurement approach provides sufficient information for fluorescence lifetime determination in imaging applications while dramatically reducing the data acquisition and processing time required for full spectroscopic characterization
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 enables accurate and precise measurement of fluorescence lifetime at high speeds, suitable for FLIM microscopy, allowing for real-time image acquisition and overcoming the limitations of traditional methods by simplifying the calculation process and improving measurement speed.
Implementation Method 1
a light sensor that converts the fluorescence photons into a fluorescence electrical signal
Data Source
AI summary
Disclosed is an apparatus for measuring a fluorescence lifetime. The apparatus for measuring the fluorescence lifetime comprises an excitation light generator that generates excitation light to be irradiated on a sample including fluorescence molecules; a fluorescence photon collecting unit that collects a plurality of fluorescence photons generated by irradiating the excitation light on the sample; a light sensor that converts the collected fluorescence photons into a fluorescence electrical signal; and a fluorescence lifetime signal processor that determines the fluorescence lifetime by calculating the average time of the fluorescence electrical signal with respect to a predetermined apparatus delay time. According to the above configuration, the present invention can accurately and precisely measure a fluorescence lifetime in a short measurement time by easy calculation.