Fluorescence-lifetime measurements in atmospheric-pressure flames using nanosecond-pulsed lasers

Measurements of fluorescence lifetimes are needed to quantify concentration measurements when using linear laser-induced fluorescence. However, lifetimes are only a few nanoseconds for many important species at atmospheric pressure. When using a typical Q-switched laser with a pulse width of about 10 ns, the fluorescence follows the shape of the laser pulse and the lifetime cannot be easily measured. In this paper, a technique is described for experimentally determining the fluorescence lifetime in atmospheric-pressure flames using a nanosecond-pulsed laser; that is, measurement of a lifetime an order-of-magnitude faster than the laser pulse itself. This technique relies on an observable temporal shift in the fluorescence signal as a function of the lifetime. Simulations show the efficacy of this approach, and data in liquid samples and in an atmospheric-pressure flame show excellent agreement with prior picosecond measurements. This technique is successful because only the temporal shift is examined and details of the fluorescence profile are ignored. Received: 23 August 2001 / Revised version: 19 November 2001 / Published online: 17 January 2002     

激光诱导叶绿素荧光寿命的测量及其特性分析

提出了一种用于评估植物生长状况及环境监测的激光诱导叶绿素荧光寿命测量方法. 采用波长355 nm的激光作为光源激发叶绿素荧光, 由光电倍增管接收其荧光信号, 由于被测叶绿素荧光衰减函数与激光脉冲、仪器响应函数卷积在一起, 根据它们的特性, 运用时间分辨测量法分别测得叶绿素荧光及其背景信号, 并结合一种新型解卷积算法可分离出真实的叶绿素荧光衰减函数, 从而获取叶绿素的荧光寿命. 测试结果表明: 该方法能够实现叶绿素荧光寿命的高精度实时监测, 对不同叶绿素含量的溶液荧光寿命进行了测试, 证明叶绿素含量与其荧光寿命具有相关性, 并且拟合了叶绿素含量与荧光寿命的标定曲线. 关键词： 荧光寿命 激光诱导荧光 时间分辨测量法 叶绿素含量     

基于激光诱导叶绿素荧光寿命成像技术的植物荧光特性研究

针对植物荧光遥感探测中信号易受干扰的问题, 提出了一种用于评估植物生长状况及环境监测的荧光寿命成像技术. 采用凹透镜对355 nm波长的激光扩束, 再照射植物激发叶绿素荧光, 由增强型电荷耦合器件接收荧光信号. 采用时间分辨测量法, 连续用相同激光脉冲照射植物以激发相同的荧光信号, 同时不断改变激光脉冲触发探测器启动的延时时间, 从而能够得到完整的离散荧光信号分布图像. 对植物特定位置点产生的离散荧光信号进行拟合, 再运用一种改进型的迭代解卷积法可反演高精度的荧光寿命; 进而反演图像各点的荧光寿命以生成植物的荧光寿命分布图. 该方法所绘制的荧光寿命图比荧光强度图能更准确地反映植物内部的叶绿素含量, 并对活体植物叶绿素荧光寿命的物理特性进行了初步研究, 证明叶绿素荧光寿命与植物生理状态存在一定关联; 并且叶绿素荧光寿命与活体植物所处环境存在着复杂的关系. 未来将与生物物理学家们合作, 继续探寻叶绿素荧光寿命与植物生存环境的关系.     

Confocal fluorescence lifetime imaging of free calcium in single cells

Ca²⁺ concentrations in biological cells are widely studied with fluorescent probes. The probes have a high selectivity for free calcium and exhibit marked changes in their photophysical properties upon binding. The differences in the fluorescent lifetime of the probes can now be used as a contrast mechanism for imaging purposes. This technique can be further exploited for the quantitative determination of ion concentrations within the cells. We describe the use of a fast fluorescence lifetime imaging method in combination with a standard confocal laser scanning microscope for the determination of Ca²⁺ concentrations in single rat cardiac myocytes using the intensity probe Calcium Green.     

Whole-field five-dimensional fluorescence microscopy combining lifetime and spectral resolution with optical sectioning

We report a novel whole-field three-dimensional fluorescence lifetime imaging microscope that incoporates multispectral imaging to provide five-dimensional (5-D) fluorescence microscopy. This instrument, which can acquire a 5-D data set in less than a minute, is based on potentially compact and inexpensive diode-pumped solid-state laser technology. We demonstrate that spectral discrimination as well as optical sectioning minimize artifacts in lifetime determination and illustrate how spectral discrimination improves the lifetime contrast of biological tissue.     

Kinetics of Er3+-doped fluorozirconate optical fiber upconversion fluorescence and laser emission under 800 nm excitation

Applying single- and double-pulse excitation at 800 nm, the kinetics of the upconversion fluorescence in the green, as well as the upconversion laser at 543 nm was studied. No significant delay between the pumping pulse and the laser emission was found. In the erbium doped (1000 ppm) optical fiber, the mechanism responsible for the upconversion is purely of the excited state absorption (ESA) type. The double-pulse technique enables also a determination of the lifetime of the intermediate metastable state ⁴ I _11/2 (7±0.3 ms). Some other basic properties of the upconverted fluorescence and of the laser itself (fluorescence spectrum, optical gain, laser threshold) are also described. Received: 11 December 2000 / Revised version: 18 February 2001 / Published online: 18 July 2001     

Photon-counting technique for rapid fluorescence-decay measurement

We report on a novel laser-induced fluorescence triple-integration method (LIFTIME) that is capable of making rapid, continuous fluorescence lifetime measurements by a unique photon-counting technique. The LIFTIME has been convolved with picosecond time-resolved laser-induced fluorescence, which employs a high-repetition-rate mode-locked laser, permitting the eventual monitoring of instantaneous species concentrations in turbulent flames. We verify the technique by application of the LIFTIME to two known fluorescence media, diphenyloxazole (PPO) and quinine sulfate monohydrate (QSM). PPO has a fluorescence lifetime of 1.28 ns, whereas QSM has a fluorescence lifetime that can be varied from 1.0 to 3.0 ns. From these liquid samples we demonstrate that fluorescence lifetime can currently be monitored at a sampling rate of up to 500 Hz with less than 10% uncertainty (1sigma) .     

Enhancement of rapid lifetime determination for time-resolved fluorescence imaging in forensic examination

An enhancement method of rapid lifetime determination is proposed for time-resolved fluorescence imaging to discriminate substances with approximate fluorescence lifetime in forensic examination. In the method, an image-exclusive-OR treatment with filter threshold adaptively chosen is presented to extract the region of interest from dual-gated fluorescence intensity images, and then the fluorescence lifetime image is reconstructed based on the rapid lifetime determination algorithm. Furthermore, a maximum and minimum threshold filtering is developed to automatically realize visualization enhancement of the lifetime image. In proof experiments, compared with traditional fluorescence intensity imaging and rapid lifetime determination method, the proposed method automatically distinguishes altered and obliterated documents written by two brands of highlighters with the same color and close fluorescence lifetime.     

Quantitative imaging of mixing dynamics in microfluidic droplets using two-photon fluorescence lifetime imaging

Droplet-based microfluidic systems enable miniaturization of chemical reactions in femtoliter to picoliter volume compartments. Quantifying mixing dynamics of the reagents in droplets is critical to determine the system performance. In this Letter, we developed a two-photon excitation fluorescence lifetime imaging technique to quantitatively image the mixing dynamics in micro?uidic droplets. A cross/autocorrelation method was used to reconstruct a high-quality fluorescence lifetime image of the droplet. The fluorescence decay was analyzed for accurate determination of the mixing ratio at each pixel of the image.     

Measurement of upper level lifetime in presence of radiation trapping: A revisitation of the pinhole method

In this work we expose a recently developed theoretical model for the analysis of the measurements obtained with the so-called pinhole method, for the determination of the upper level lifetime in materials where the superimposition between fluorescence and absorption spectra determines significant radiation trapping effect. Under fairly general conditions the fluorescence decay curve after a pulsed excitation of the sample obtained with this experimental set-up can be conveniently described by a double exponential functional form. The faster of the two decay times is close to the intrinsic fluorescence lifetime, with a difference that can be calculated with geometrical considerations. The theoretical results were tested with a suitably designed experiment, where the upper level lifetime of the laser transition of some Yb doped samples were investigated, obtaining results that are consistent with literature data and in good agreement with the theoretical results as for the temporal dependence of the fluorescence decay.     

Photo-physical and lasing characterisation of a polyparaphenylenevinylene (PPV) neat film

The wave-guided travelling-wave laser action (amplified spontaneous emission) of a neat film of poly(p-phenylenevinylene) (PPV) on a quartz glass substrate prepared by a sulfinyl precursor technique is studied. The samples are transversally pumped with picosecond excitation pulses (wavelength 347.15 nm, duration 35 ps). Lasing occurs at 550 nm. The optical constants of the neat films are determined by transmittance measurements exploiting the multiple beam interference in the transparency region. A fluorescence spectroscopic characterisation is carried out determining the fluorescence quantum distribution, fluorescence quantum yield, degree of fluorescence polarisation, and fluorescence lifetime. The emitting chromophore size (emitting singlet exciton extension) is determined by the ratio of exciton radiative lifetime to repeat-unit based radiative lifetime. The obtained size of about two repeat units is discussed in a disordered solid-state polymer model.     

基于超短脉冲的癌细胞荧光光谱

研究用于癌症诊断与治疗的光敏剂血卟啉(hematoporphyrin derivative,HPD)的超快光动力学过程。采用超短脉冲激光光谱技术和皮秒时间相关单光子计数系统,测量经血卟啉培养的活体癌细胞与正常细胞的荧光光谱、荧光寿命特性及荧光峰值强度随时间的变化,观测到:癌细胞样品在645nm处具有特征发射光谱峰;癌细胞与正常细胞样品荧光寿命的快成分分别为150,300ps;癌细胞与正常细胞的荧光峰值强度经12h分别衰减10%和55%。对测量所得的荧光光谱曲线及时间分辨荧光衰减曲线分析,计算出:在癌细胞内部血卟啉浓度增大了约2个数量级;癌细胞与正常细胞的荧光寿命分别为824,1798ps;血卟啉在癌细胞与正常细胞样品中滞留时间分别为17,6d。测量结果确认了荧光光谱技术诊断与治疗癌症的可行性,并对发展超短脉冲激光光谱技术早期诊断与治疗癌症具有重要的指导意义和临床应用价值。     

基于不同泵浦波形的荧光寿命测量

该文从理论上分析了不同泵浦波形下的荧光衰减规律,提出了一种在不同泵浦波形下测量荧光寿命的新方法——双脉冲探测法,即利用探测泵浦脉冲与荧光衰减脉冲的方法测量荧光寿命。通过对样品钕玻璃及Cr:ZnSe晶体的荧光寿命测量表明:利用该方法在不同泵浦波形下能够实现可见及近红外到中红外激光介质的荧光寿命测量。因此,利用该测量方法能够方便、有效的避免通过解卷积求样品荧光寿命的繁琐过程,对测量激光介质在不同泵浦波形下的荧光寿命具有参考价值。     

NO分子A^2∑←X^2∏跃迁的激光双光子荧光激发谱

以Nd：YAD脉冲激光器泵浦的光学参量发生器／放大器(OPG／OPA)作激发光源,获得了420～472nm波长范围内NO分子的双光子激光荧光激发谱,并利用此技术对N0分子的能级结构进行了实验研究,将所得谱线峰归属为NO(A^2∑←X^2∏)的跃迁,荧光强度随激光强度的二次方变化关系表明此过程是一双光子激发过程。利用实验所得峰值波长计算了NO(A^2∑)态的基振动频率ωe和平衡位置的力常数k。通过对NO分子A^2∑←X^2∏跃迁的荧光时间分辨光谱进行实验研究,得到266Pa气压下A^2∑(v′=0)态的能级寿命r=53．76ns。测量荧光寿命随气压的变化,利用曲线拟合得到NOA。三(v′=0,1)两振动态的自发辐射寿命和无辐射跃迁驰豫速率常数。     

Fluorescence lifetime system for microscopy and multiwell plate imaging with a blue picosecond diode laser

We report a wide-field fluorescence lifetime imaging (FLIM) system that uses a blue picosecond pulsed diode laser as the excitation source. This represents a significant miniaturization and simplification compared with other time-domain FLIM instruments that should accelerate the development of clinical and real-world applications of FLIM. We have demonstrated this instrument in two configurations: a macroimaging setup applied to multiwell plate assays of chemically and biologically interesting fluorophores and a microscope system that has been applied to imaging of tissue sections. The importance of the adjustable repetition rate of this laser source is discussed with respect to noise reduction and precision in the lifetime determination, illustrating a further significant advantage over conventional mode-locked solid-state lasers.     

钇原子亚稳态寿命的测量

利用基态原子无穷长的寿命以及亚稳态原子在飞行过程中的衰减,比较由激光感应荧光法测量到的钇原子束基态和亚稳态的速度分布曲线,经过拟合处理计算,得到钇原子亚稳态a~2D_(5/2)的寿命为(0.8±0.2)ms.     

高浓度若丹明6G溶液荧光寿命的测量

本文提出一种测量高浓度溶液荧光寿命的方法.利用这种方法测量了浓度范围从1×10-6至1×10-2mol·L-1的Rh6G乙醇溶液的荧光寿命.并证明了在这个浓度范围内,Stern-Volmer公式仍然成立.     

激光诱导植物荧光寿命校正技术及其特性分析

激光诱导植物荧光寿命测量法是在植物荧光光谱分析法基础上开发的一种评估植物生长状况及环境监测的新技术。根据植物叶绿素荧光信号的物理特性,利用信息仿真技术开发了一种叶绿素荧光寿命校正方法,可提高植物叶绿素荧光寿命的测量精度。利用激光诱导叶绿素荧光寿命测量系统分别测得叶绿素荧光及其背景信号,先用解卷积法叶绿素荧光信号中分离出荧光衰减函数,可获取荧光寿命估计值。再结合叶绿素荧光寿命校正技术就能反演得到高精度的植物荧光寿命。仿真与实验结果表明：该方法可实现高精度的植物荧光寿命实时监测;并对不同含量的叶绿素提取液进行了测试,构建了植物荧光寿命与叶绿素含量的对应关系模型。未来该技术可用于遥感监测海洋、湖泊、河流中藻类植物的生物含量。     

通过光致还原调制氧化石墨烯寿命并用于微纳图形制备

氧化石墨烯因其宽带可调谐的荧光发射特性已被广泛应用于荧光成像、金属离子高灵敏检测和光电器件的制备.相比于荧光强度,氧化石墨烯荧光寿命不受材料厚度和激发功率的影响,具有更为稳定和均一的特性.本文研究了在激光还原过程中氧化石墨烯荧光寿命逐渐减小的变化行为,发现了长寿命sp~3杂化结构向短寿命sp~2杂化结构的转变.通过精确控制还原时间,结合激光直写技术,在单层氧化石墨烯薄膜上实现了二维码、条形码、图形和数字等微纳图形的制备,还在多层氧化石墨烯薄膜结构上获得了多寿命多层微纳图形.这种微纳图形的制备具有灵活无掩膜、高对比和多模式的特点,可用于高密度光学存储、信息显示和光电器件制备等诸多领域.