Time-resolved fluorescence intensity issued from a heterogeneous slab: Sensitivity characterization

Optical imaging using fluorescent contrast agents has become an interesting tool to differentiate diseased lesions from normal tissue. However, several sensitivity characterizations may strongly influence the time-dependent fluorescence measurements. Herein, we present a numerical model based on the finite element method that allows the simulation of time-resolved reflectance and transmittance signals from heterogeneous media mimicking breast tissues with an embedded fluorescent object (tumor). The influence, on the computed signals, of several tumor depths, as well as various fluorophore concentrations and several fluorescent markers targeting are analyzed. The results show the possibility of uncoupling location depth from the shape of the target. Therefore, the analysis of the time to reach half the maximum intensity is validated as a good localization scheme. Then, the transmitted data show that the maximal detected intensity at the bottom of the medium is very sensitive to the dye concentration but not to the tumor shape. Moreover, the strong competition between concentration determination and fluorophore distribution is presented. These results will lead to a better detection and localization of tumors.     

Refocusing and locating effect of fluorescence scattering field

Optical imaging deep inside scattering medium has always been one of the challenges in the field of bioimaging, which significantly drawbacks the employment of con-focal microscopy system. Although a variety of feedback techniques, such as acoustic or nonlinear fluorescence-based schemes have realized the refocusing of the coherent light, the problems of non-invasively refocusing and locating of linearly-excited fluorescent beads inside the scattering medium have not been thoroughly explored. In this paper, we linearly excited the fluorescent beads inside a scattering medium by using our homemade optical con-focal system, collected the fluorescence scattering light as the optimized target, and established a theoretical model of target contrast enhancement, which is consistent with the experimental data. By improving both the cost function and variation rate within the genetic algorithm, we could refocus the fluorescence scattering field while improving the contrast enhancement factor to 12.8 dB. Then, the positions of the fluorescent beads are reconstructed by sub-pixel accuracy centroid localization algorithm, and the corresponding error is no more than 4.2 μ with several fluorescent beads within the field of view. Finally, the main factors such as the number of fluorescent beads, the thickness of the scattering medium, the modulating parameter, the experimental noise and the system long-term stability are analyzed and discussed in detail. This study proves the feasibility of reconstructing fluorescent labeled cells inside biological tissues, which provides certain reference value for deep imaging of biological tissues.     

Laser-Induced Fluorescence Bronchoscopy for Detection and Localization of Early Lung Cancer

Experimenatal results on the development of a Laser-Induced Fluorescence Bronchoscopy(LIFB) for the detection and localization of early lung cancer are reported in this paper. The system utilizes fluorescence of photosensitizer drug to provide real time video imaging for the examined lung tissue. Color filters are used to differentiate signal from background and a computer image processing technique is also applied to subtract the background. Moreover, a pseudocolor contrast enhancement method was developed to enhance the fluorescence image displayed on the vidio monitor. Suspicious areas are identified by pseudocolor image to guide biopsy, and several clinical trials show that sensitivity and contrast capability of the system should permit the detection and localization of early lung cancer.     

Laser-Induced Fluorescence Bronchoscopy for Detection and Localization of Early Lung Cancer

Experimenatal results on the development of a Laser-Induced Fluorescence Bronchoscopy(LIFB) for the detection and localization of early lung cancer are reported in this paper. The system utilizes fluorescence of photosensitizer drug to provide real time video imaging for the examined lung tissue. Color filters are used to differentiate signal from background and a computer image processing technique is also applied to subtract the background. Moreover, a pseudocolor contrast enhancement method was developed to enhance the fluorescence image displayed on the vidio monitor. Suspicious areas are identified by pseudocolor image to guide biopsy, and several clinical trials show that sensitivity and contrast capability of the system should permit the detection and localization of early lung cancer.     

具有图象处理功能的激光荧光内窥系统的研究

内窥镜荧光图象系统是体腔内早期肿瘤诊断和定位的有效手段.但是早期开发研究的LFE荧光成家系统由于存在假阳性和假阴性误诊而限制了它的广泛开展和应用.本文仔细探讨了该系统产生误诊的原因,并在此基础上提出了使用计算机图象处理技术的荧光图象系统,而且通过实验验证了这种新技术,它能有效地克服假阳性和假阴性误诊,为体腔内肿瘤的诊断提供可靠判据.     

高精度实时主动轴向防漂移系统研究

基于单分子定位的荧光纳米分辨显微成像中, 系统漂移会使得单分子定位出现额外偏差, 从而使重构图像的分辨率降低, 造成图像模糊. 因此, 对系统漂移量的控制至关重要. 近年来, 防漂移的方法层出不穷. 本文针对其中一种利用光学测量原理和引入负反馈的防漂移方法做了系统的研究, 分析了其原理和实现过程, 对整个系统进行了误差分析, 通过实验标定了整个防漂移系统的精度. 该系统可以主动实时地校正漂移量, 实现了显微镜轴向9.93 nm的防漂移精度. 与现有商用的显微镜自带的防漂移装置相比, 防漂移精度提高了一个量级.     

基于差分光密度差异的组织内异质体的快速定位方法

基于近红外光谱法对组织内的异质体进行无创检测时,光源-探测器(S-D)相对于异质体的位置对检测效果有着重要影响。为实现对组织内异质体的快速定位,该研究基于一源多探的检测结构针对不同水平位置、不同深度和不同直径的异质体进行光密度分布有限元分析,计算各探测器之间的差分光密度差异。仿真实验结果表明,根据多探测器形成的差分光密度差异曲线可快速定位组织内异质体的水平位置。曲线的高斯拟合特征量与异质体的水平位置、深度和直径有着强相关性。基于差分光密度差异曲线可以实现组织内感兴趣区域的快速定位,对采用近红外光谱法的组织肿瘤检测、光学脑功能成像等领域的源-探位置放置提供重要参考,提高其检测精度。     

DNA microarray scanner with a DVD pick-up head

A low-cost highly sensitive DNA microarray scanner for fluorescent detection is developed based on the pick-up head of a commercially available optical storage device, DVD. A laser beam of 650 nm, generated by a DVD laser diode, is used for dynamic auto-focusing as well as the excitation of Cy5 fluorescent dye. The fluorescence intensity emitted from Cy5 dye is measured by a photomultiplier tube (PMT). In contrast to other microarray scanners, the DVD-based scanner offers the auto-focusing function using the focus error signal (FES) and a voice coil motor (VCM), and this enables fast response, high accuracy and compact size. The fluorescence-detecting performance of the scanner is inspected by using a commercial BAC (bacterial artificial chromosome) oligonucleotide chip and a scanner evaluation microarray (DS01). Experiments have shown that the DVD-based scanner meets the limit of detection, ensuring the feasibility of a low-cost, highly sensitive DNA microarray scanner.     

Fast Fourier domain localization algorithm of a single molecule with nanometer precision

We present an algorithm to determine the location of a fluorescent molecule with nanometer-scale accuracy. A Fourier domain localization scheme based on zero-padded fast Fourier transform and phase gradient operators is used to obtain a powerful mathematical model for localizing the molecule without numerical fitting. Compared with conventional algorithms, our position estimator does not require prior background information or initial parameter estimation. Numerical simulations indicate that the proposed method exhibits high localization precision and small bias while executing almost as fast as the fluoroBancroft algorithm.     

Fluorescent Biological Label for Recognizing Human Ovarian Tumor Cells Based on Fluorescent Nanoparticles

In this paper, we report a method for recognizing human ovarian tumor(HOT) cells using fluorescent biological label based on core-shell nanoparticles. The luminescent nanoparticles were synthesized with a water-in-oil(W/O)micromulsion technique. The fluorescent silica core-shell nanoparticles modified with anti-HER2 antibody using bifunctional cross-linker glutaraldedhyde targeted the corresponding tumor antigen in the cell surface of the SKOV3 ovarian cancer cells. The specific immunoreactivity of antibody-nanoparticles with cells was characterized by laser scanning microscopy (LSM) and scanning electron microscope (SEM). The results showed that the method offered potential advantages of sensitivity and simplicity due to high binding efficiency between nanoparticles and cells and provided an alternative method for the detection of HOT.     

Axial coding in full-field microscopy using three-dimensional structured illumination implemented with no moving parts

We report a simple optical setup to produce both axial and lateral structured illumination through a single objective lens. With a minimum of six full-field images obtained without moving either the sample or the microscope objective, 100 nm diameter fluorescent beads can be localized axially with an accuracy of 50 nm in a 1.76-microm-thick layer. We show that this axial localization improvement can easily be combined with classical lateral structured illumination, so that lateral resolution enhancement by a factor of 2 is maintained.     

磁免疫和荧光免疫传感器的构建及其在检测甲胎蛋白中的应用

甲胎蛋白(AFP)是常见的肝癌肿瘤标志物,在早期诊断方面起到重要作用。分别设计构建了磁免疫和荧光免疫传感器并将其应用于AFP的定量检测。在磁免疫传感器中,采用免疫磁珠代替传统固相载体,实现了目标物的快速分离;利用标记抗体上的辣根过氧化物酶(HRP)催化底物显色,根据底物吸光度值的大小进行定量检测。构建的AFP检测方法的检出限为3.6 ng·mL-1,线性范围为10~80 ng·mL-1。在荧光免疫传感器中,将碲化镉量子点(CdTe QDs)的荧光作为信号输出,并通过同时将多个CdTe QDs连接在纳米硅球表面实现信号放大,通过测量量子点荧光强度实现定量检测。该方法AFP检出限为4.2 ng·mL-1,线性范围为5~150 ng·mL-1。所设计的两种传感器均具有特异性强、灵敏度高的特点,为AFP的检测提供了新的思路和方法。     

IMPROVED DAMAGE IDENTIFICATION METHOD BASED ON MODAL INFORMATION

In this paper, a newly derived algorithm to predict locations and severities of damage in structures using changes in modal characteristics is presented. First, two existing algorithms of damage detection are reviewed and the new algorithm is formulated in order to improve the accuracy of damage localization and severity estimation by eliminating erratic assumptions and limits in the existing algorithms. Next, the damage prediction accuracy is numerically assessed for each algorithm when applied to a two-span continuous beam for which pre- and post-damage modal parameters are available for only a few modes of vibration. Compared to the existing damage detection algorithms, the new algorithm improved the accuracy of damage localization and severity estimation results in the test beam.     

肿瘤细胞中表达的GFP蛋白的荧光漂白特性的研究

GFP作为生物源性荧光探针具有其他荧光标记物所无法比拟的优势，目前已广泛应用于生物学研究的各个领域。利用常规转染方法将带有EGFP基因的质粒载体导入人肺腺癌肿瘤细胞(ASTC-a-1)，并得到GFP稳定表达的细胞株。研究中发现，肿瘤细胞中表达的GFP长时间暴露于强激发光中会发生非常强列的荧光漂白作用，并且这种漂白作用是不可恢复的。对不同强度的激发光(饱和光源、阻断片ND4／ND8／ND16)对GFP的漂白作用进行了研究，并对冷冻保存样品的光漂白作用进行了初步的探讨。     

基于改进的Mask R-CNN的乳腺肿瘤目标检测研究

乳腺癌是全球女性死亡率最高的恶性肿瘤之一,早期发现有助于提升患者的存活率。本文利用深度学习中的目标检测网络对乳腺X线图像中的肿瘤病变区域进行定位和分类;然后选取Mask R-CNN网络作为目标检测模型,对Mask R-CNN的基准网络D-ShuffleNet进行改进,提出了一种新的网络——Mask R-CNN-II网络,并在Mask R-CNN-II网络中应用迁移学习算法。通过实验验证了Mask R-CNN-II网络比Mask R-CNN网络的检测精度更高,而且验证了所提基准网络、所使用的融合图像的思想以及迁移学习算法是有效的。Mask R-CNN-II有利于提高乳腺肿瘤的定位与分类,可为放射科医生提供辅助诊断意见,具有一定的临床应用价值。     

基于荧光光谱法与深度信念网络的稻种发芽率检测方法研究

针对传统稻种发芽率检测效率低、精度差、专业化要求高等问题,通过荧光光谱法结合深度信念网络(DBN)建立稻种发芽率预测模型。首先,将连粳7号和武运粳均分别老化0~7 d后,以5 min为间隔在纯净水中分别浸泡5~30 min。然后用荧光光谱仪检测浸泡液的荧光光谱,光谱数据经中心化后用集合经验模态分解(EEMD)去噪,并通过主成分分析法提取441.5 nm的特征荧光波长。最后,利用偏最小二乘回归(PLSR),反向传播神经网络(BPNN),径向基函数神经网络(RBFNN)和深度信念网络(DBN)建立水稻种子发芽预测模型。比较后得出,DBN模型在少数据、弱信号情况下的预测精度最高,预测集相关系数Rp和均方根误差RMSEP最大可达0.979 2和0.101。同时,通过分析混合稻种荧光数据R_p的变化趋势,得到最佳浸泡时间为22.1 min,实际上,精确度超过0.95(R_p)需要5 min左右。研究结果表明,结合荧光光谱法和EEMD-DBN模型,非破坏性地预测水稻种子发芽率具有可行性和高准确性,并且适用于不同颜色和污染水平的水稻种子的检测。     

Local fluorescence spectroscopy and detection of malignancies using laser excitation at various wavelengths

Comparative study of the sensitivity of the local fluorescence spectroscopy in the detection of malignancies with a size of 1–7 mm is performed for various fluorescence excitation wavelengths of the injected photosensitizing agent. Several algorithms for the calculation of the fluorescence contrast of malignant lesion against the adjacent uninvolved tissue are analyzed using the Sarcoma 37 experimental tumor model. The photosensitizers from the Porphyrin (e.g. the 5-ALA-induced Protoporphyrin IX) and Chlorin (e.g. Chlorin e6) families are selected as the fluorescent marker of tissues. Three laser sources with radiation wavelengths of 408, 532, and 635 nm which fall in the absorption bands of Porphyrins and Chlorines are used for the tissue fluorescence excitation.     

Detecting enzymes in living cells using fluorogenic substrates

Characteristics of fluorogenic substrates designed for detection of enzyme activity in living cells are reviewed. Improved retention of the fluorescent products in the cell of origin can be achieved by structural modifications to the substrate that result in association with membrane lipids or conjugation to intracellular glutathione. Newly-developed substrates that yield fluorescent precipitates provide the additional advantage of allowing subcellular localization of sites of enzymatic activity. Improved detection sensitivity can also be achieved by targeted delivery of substrates for processing by specific organelles. Substrates designed for monitoring oxidative activity and lipid metabolism provide examples of this approach.     

HeLa细胞突起中微丝束的纳米分辨荧光成像

在Matlab编程环境下模拟了单分子定位显微的纳米分辨成像,在传统实验参数条件下,对不同间隔分子带模型进行了模拟成像.模拟结果表明,单分子定位显微方法可以区分中心相隔20 nm的两个分子带.同时,也分析了不同像元大小对单分子定位精度的影响.此外,通过单分子定位显微方法在IX71倒置荧光显微镜上实现了纳米分辨,系统极限分辨率,即半高全宽为48 nm.在该系统上,获得了HeLa细胞突起中微丝束结构的纳米分辨图像.从重构获得的图像中可以看到微丝束的直径为75—200 nm.     

氢化物发生-原子荧光光谱法测定化妆品中的铅

本文介绍了氢化物发生－原子荧光光谱法对化妆品中铅含量的定量分析方法,并研究拟定了原子荧光光谱法测定铅的分析条件,本方法回收率为96.0%－101.5%,检出限为0.18ng/mL,精密度（RSD）为0.90%-0.94%,准确性好,可用于测定化妆品中铅的含量。