Fluorescence decay data analysis correcting for detector pulse pile-up at very high count rates
Fluorescence decay data analysis correcting for detector pulse pile-up at very high count rates
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DOI：
10.1117/1.oe.57.3.031305
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发表时间：
2018-03-01
影响因子：
1.3
通讯作者：
Wahl, Michael
中科院分区：
文献类型：
--
作者：
Patting, Matthias;Reisch, Paja;Wahl, Michael
使用与时间相关的单光子计数用于荧光寿命测量的目的通常由于堆积而速度受限。利用现代仪器，这一限制可以被显著地解除，但由于紧密间隔的探测器脉冲(探测器脉冲堆积)的频繁合并而导致的一些伪影仍然是需要解决的问题。我们提出了一种数据分析方法，对这种伪影和由此产生的系统误差进行校正。它对探测器脉冲堆积引起的光子损失进行物理建模，并将该损失并入衰变拟合模型中，以获得衰变分量的荧光寿命和相对幅度。对有和没有这种修正的结果的比较表明，在接近激励速率的计数速率下，系统误差显著减小。这允许在非常高的帧速率下进行定量准确的荧光寿命成像。(C)2018年光学仪器工程师学会(SPIE)。
Using time-correlated single photon counting for the purpose of fluorescence lifetime measurements is usually limited in speed due to pile-up. With modern instrumentation, this limitation can be lifted significantly, but some artifacts due to frequent merging of closely spaced detector pulses (detector pulse pile-up) remain an issue to be addressed. We propose a data analysis method correcting for this type of artifact and the resulting systematic errors. It physically models the photon losses due to detector pulse pile-up and incorporates the loss in the decay fit model employed to obtain fluorescence lifetimes and relative amplitudes of the decay components. Comparison of results with and without this correction shows a significant reduction of systematic errors at count rates approaching the excitation rate. This allows quantitatively accurate fluorescence lifetime imaging at very high frame rates. (c) 2018 Society of Photo-Optical Instrumentation Engineers (SPIE).