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Method and device for measuring fluorescence lifetime

A fluorescence lifetime and measurement method technology, applied in fluorescence/phosphorescence, material excitation analysis, etc., can solve the problems of high price, time-consuming adjustment and processing, complex optical path, etc., and achieve low-cost, fast and high-throughput real-time online measurement , the effect of simple installation

Active Publication Date: 2019-06-11
ZHEJIANG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0003] However, both of these technologies must use pulsed laser light as a light source, with a photomultiplier tube (or avalanche photodiode) and an enhanced charge-coupled device camera as a detector, which is very expensive
In addition, these two methods need to drive the detection device regularly according to the irradiation of the excitation light source, the optical path is complicated, and the adjustment and processing are time-consuming.

Method used

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  • Method and device for measuring fluorescence lifetime
  • Method and device for measuring fluorescence lifetime
  • Method and device for measuring fluorescence lifetime

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0029] Fix the substrate with No. 1 fluorescent substance to be tested on the sample holder, and irradiate the sample with a pulse light source with a pulse repetition frequency of 1 kHz. The high-speed camera continuously shoots the fluorescent signal, and the exposure time of each frame is 50us. In the series of photos taken, a fluorescent signal corresponding to the repetition frequency of the excitation pulse appears, and the fluorescent signal changes periodically from strong to weak. A total of 20 photos in which the fluorescent signal continuously changes from strong to weak were randomly selected. Capture the corresponding position of the fluorescent substance in the photo, such as figure 1 shown. Then plot the sum of the gray values ​​of all pixels in each frame against time to obtain the fluorescence decay curve, as shown in figure 2 shown.

[0030] Then use the single exponential decay function I(t)=I bg +A·exp(-t / τ) fits the data points, where, I bg is the b...

Embodiment 2

[0032] Fix the substrate with No. 2 fluorescent substance to be tested on the sample holder, and irradiate the sample with a pulse light source with a pulse repetition frequency of 1 kHz. The high-speed camera continuously shoots the fluorescent signal, and the exposure time of each frame is 50us. In the series of photos taken, a fluorescent signal corresponding to the repetition frequency of the excitation pulse appears, and the fluorescent signal changes periodically from strong to weak. A total of 20 photos in which the fluorescent signal continuously changes from strong to weak were randomly selected. Capture the corresponding position of the fluorescent substance in the photo, such as image 3 shown. Then plot the sum of the gray values ​​of all pixels in each frame against time to obtain the fluorescence decay curve, as shown in Figure 4 shown.

[0033] First use the single exponential decay function I(t)=I bg +A·exp(-t / τ) fits the data points, where, I bg is the ...

Embodiment 3

[0035] Fix the substrate with No. 3 fluorescent substance to be tested on the sample holder, irradiate the sample with a pulse light source, and operate according to the method of Example 1 (the exposure time of each frame is 100 μs, continuous shooting pulse light source excitation, repetition frequency is 500μs), get as Figure 5 Fluorescence decay images are shown. Due to the high pulse repetition frequency of the excitation light source, the fluorescence signal of the substance to be measured has not decayed to a lower value, and the next excitation pulse has arrived.

[0036] In this case, multi-segment fluorescence decay time series can be used for fitting, and the fitting result is as follows Image 6 As shown, using the calculation formula in Example 2, the average value of the obtained fluorescence lifetime is 200 μs.

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Abstract

A fluorescence lifetime measurement method and device. The device comprises a sample-containing apparatus, an excitation apparatus, an imaging apparatus and a data processing apparatus. The method comprises: placing a fluorescence sample on the sample apparatus; using a light source of the excitation apparatus to excite the fluorescence of the sample; then turning off the light source and using the imaging apparatus to image the fluorescence emitted by the sample in real-time, so as to obtain a series of images of fluorescence intensity as a function of time; and using the series of images to work out the fluorescence lifetime. The device is simple, has low cost and is easy to operate, which can realize quick and high-throughput real-time online measurement.

Description

technical field [0001] The invention relates to a fluorescence lifetime measurement method and device, which are used to detect the technical field of lifetime measurement and imaging of emitted light caused by excitation light irradiation. Background technique [0002] At present, there are two main methods for the measurement and imaging of fluorescence lifetime: one is to use time-correlated single photon counting (TCSPC) technology to measure the fluorescence lifetime, which uses photomultiplier tube or avalanche photodiode as scanning method for fluorescence lifetime imaging. The second is to use time-gated technology, using an enhanced charge-coupled device (ICCD) camera as a detector, and collect fluorescence signals with a delay of a certain time after each light pulse is excited, and repeat this process many times with different time delays. The fluorescence lifetime is measured, and then combined with the wide-field imaging method, the fluorescence lifetime imagin...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N21/64
CPCG01N21/64
Inventor 秦海燕彭笑刚
Owner ZHEJIANG UNIV