TRF检测法在核酸分析中的应用

综述了镧系离子时间分辨荧光检测法在核酸分析技术中的应用概况。介绍了ＤＥＬＦＩＡ系统、ＦＩＡｇｅｎ系统、ＥＡＬＬ系统和ＴＢＰ系统的使用特点，包括时间分辨荧光分析原理、镧系离子标记ＤＮＡ探针和运用时间分辨荧光技术检测ＰＣＲ产物的技术和方法。     

TRF检测法在核酸分析中的应用

综述了镧系离子时间分辨荧光检测法在核酸分析技术中的应用概况。介绍了ＤＥＬＦＩＡ系统、ＦＩＡｇｅｎ系统、ＥＡＬＬ系统和ＴＢＰ系统的使用特点，包括时间分辨荧光分析原理、镧系离子标记ＤＮＡ探针和运用时间分辨荧光技术检测ＰＣＲ产物的技术和方法     

铽在生物活性分析中的应用

介绍了铽（Ｔｂ３＋）的理化性质、发光机理及其配体。综述了近年来Ｔｂ３＋在酶放大镧系发光法（ＥＡＬＬ）、多标记的时间分辨荧光免疫分析（ＴＲＦＩＡ）、直接固相ＴＲＦＩＡ、核酸分子杂交分析、生物大分子结构检测等方面的应用。     

中国光子生物学研究与光诊断技术进展

激光技术由于具有非接触、凝血、消毒等优点而被广泛地应用到医学临床各科中，光子所特有高能量、高的空间、时间和光谱分辨的特性使得光学成像、光声成像、荧光光谱分析等光学诊断技术及相关基础研究工作得到越来越深入的发展，特别是结合现代信息检测与分析技术的高速发展，取得了诸多有益的成果。在此，简要叙述生物光子的组织内传输与相互作用分析、光学功能成像技术和光动力学疗法应用研究等方面的进展。     

高通量筛选的检测原理及应用

本综述了高通量筛选最新检测技术的原理，包括均相分析和细胞分析的检测原理及其在微量分析及高通量筛选中的应用。均相分析中应用较多的有亲合闪烁分析（SPA）、荧光分析（FA）等，而荧光分析技术主要包括荧光共振能量转移（ERT），荧光偏振（FP），时间分辨荧光（TR）以及荧光相关谱（FRS）等。细胞分析主要建立在荧光检测技术的基础上，包括第二信使分析，报告基因分析，细胞增殖分析等。高通量筛选技术的应用和发展，将会极大的促进新药研究的进程。     

时间分辨荧光免疫分析中β-NTA及其增强液的制备

目的 以自制的时间分辨荧光免疫分析中所需增强液取代昂贵的进口试剂。方法 合成增强液中主要化合物β-萘甲酰三氟丙酮,并用其配制增强液。结果 自制增强液的性能达到进口试剂水平。结论 合成路线目标产物纯度达到要求,可用于配制国产增强液。     

铕标记人血清白蛋白的研究

铕标记人血清白蛋白的研究赵启仁，林汉，张福华，刘洁在时间分辨荧光免疫分析（ＴｒＦＩＡ）技术中，制备比活性高、免疫活性强的Ｅｕ标记物是关键之一．这需要用具有双功能基团的螯合剂将Ｅｕ３＋和抗原连接起来．已有的螯合剂有：１-（对-重氮苯基）-ＥＤＴＡ、氨基苯?..     

应用荧光探针H2DCF-DA检测光照致细胞内活性氧产生的初步研究

目的应用荧光显微镜及新型荧光探针H2DCF-DA检测在光源照射过程中人脐静脉血管内皮细胞(ECV304)内活性氧的产生情况。方法将ECV304细胞接种于35mm培养皿中,24h后加入H2DCF-DA,使其终浓度为10μmol/L,孵育30min。利用荧光显微镜的激发光源作为照射光源,在采集荧光图像的同时完成光源照射,光源输出波长范围为460～490nm,功率密度约为100mW/cm2。连续采集DCF的荧光图,观察细胞内DCF荧光的变化情况,采用计算机图像处理和分析技术,求得细胞内不同照射时间点DCF平均荧光强度,进而得到平均荧光强度-时间曲线。另外,使用线粒体特异标记探针MitoFluorRed589与H2DCF-DA共同孵育细胞,分别采集同一细胞的DCF的荧光图和MitoFluorRed589的荧光图,并利用像素-荧光强度分析方法来确定DCF荧光在细胞内的分布位置。结果在光源照射的开始阶段,ECV304细胞内的DCF荧光强度迅速增加,在第10s时便上升至第2s时的2.2倍,但是随着照射时间逐渐延长,荧光强度增幅逐渐变小,到第60s时上升至第2s时的4.7倍。观察时间进一步延长,DCF的荧光强度的变化似乎进入平台期,继而开始出现下降。考察DCF荧光在ECV304细胞内的分布位置主要通过比较细胞内不同区域的I1/I2值,通过图像分析与计算得到线粒体区、细胞核区以及细胞质非线粒体区内I1/I2值分     

基于图像处理技术分析细胞吸收光敏剂过程

目的:探讨利用图像处理技术分析光敏剂进入细胞过程的可行性。方法:将光敏剂血卟啉单甲醚注入喉癌细胞的培养皿中一定时间后,利用倒置荧光显微镜在不同时刻拍摄细胞的荧光图片,应用Matlab软件等进行图像处理得到不同时刻的细胞荧光强度的参数值L,并对计算结果进行对比。     

染色体分析在辐射生物剂量学中应用的新进展

已经证明，双着丝粒畸变分析可应用于辐射生物剂量学的许多场合，如细胞遗传学与物理学方法结合可用来估算最近发生的严重过量照射。对于内科医生来说，剂量信息是很重要的，因剂量可能和随机或确定的健康风险有关。讨论了一种新近发展的用来观察稳定性易位的荧光原位杂交（ＦＩＳＨ）法及其作为回顾性剂量计应用的有关课题，如一次全身均匀照射后１１ 年获得可信的剂量估算。值得注意的是，离体实验表明，非均一或局部照射可能导致易位率随时间降低。最后对ＦＩＳＨ法所要求的工作量和其对低剂量的分辨能力的局限性进行了评论，这是在发生广泛的核素环境污染后用该方法进行人群的回顾性剂量评估所必须考虑的。     

Dyke Award. Europium-DTPA: a gadolinium analogue traceable by fluorescence microscopy

A lanthanide series chelate, europium(Eu)-DTPA, was synthesized to serve as a histochemical analogue for the widely used MR contrast agent gadolinium(Gd)-DTPA. Eu and Gd, being neighboring elements on the periodic table, share many fundamental properties, including ionic radius, valence, and chemical reactivity. Eu-DTPA, however, possesses one important physical property not shared by Gd-DTPA: luminescence under ultraviolet light. The feasibility of detecting Eu-DTPA in animal tissues under fluorescence microscopy was systematically evaluated and documented. Distinctive orange-red luminescence of Eu-DTPA could be observed in the kidneys, livers, dura, choroid, and pituitary glands of rats after intravascular injection. No luminescence was detected in areas of brain beyond an intact blood-brain barrier. When the brain was locally injured by an experimental laceration, however, leakage of Eu-DTPA was detected. Electron probe microanalysis confirmed the parallel presence or absence of simultaneously injected Eu-DTPA and Gd-DTPA in all tissues studied. Fluorescence microscopy with Eu-DTPA has thus been validated as a method for tracing the distribution of Gd-DTPA at the microscopic level.     

NMR studies of combined lanthanide shift and relaxation agents for differential characterization of 23Na in a two-compartment model system

Spin relaxation and chemical shifts by lanthanide chelate complexes are used to distinguish 23Na signals in a simulated two-compartment model. Both effects are significant in EDTA, DTPA, and TPP complexes of Gd and in the TPP complex of Dy. The simultaneous measurement of these properties is illustrated and represents a promising method for monitoring sodium concentrations and fluxes including fast transport components.     

Establishment and characterization of monoclonal antibodies against an octahedral gallium chelate suitable for immunoscintigraphy with PET.

As a prerequisite for preparing bispecific antibody conjugates containing anti-tumor and anti-metal chelate binding sites that can be used for pretargeted immunoscintigraphy, monoclonal antibodies (Mabs) have been raised against an octahedral metal chelate synthetized from gallium (Ga) and the hexadentate ligand N,N'bis[2-hydroxy 5-(ethylene beta carboxy) benzyl] ethylenediamine N,N' diacetic acid (Ga-HBED-CC). With use of the Farr assay, binding studies with the 67Ga-labeled chelate and three clones of anti-chelate Mabs showed that none of the Mabs were able to precipitate more than 50% of the Ga-chelate, suggesting an enatiomerism of the Ga-chelate and a sensitivity of the Mabs to either one or the other chelate enantiomer. This could be confirmed by comparing the circular dichroism spectra of the Ga-chelate fractions that passed affinity columns containing the Mabs immobilized on sepharose without retention. With use of a Ga-HBED-CC enantiomer, whole-body retention in mice, preinjected with the corresponding anti-metal chelate Mab of ca. 70% ID, was measured compared to 2.1% retention in mice not preinjected with the Mab. Due to the high affinity of chelate-to-Mab binding in vivo, bispecific antibody conjugates prepared from the fragments of the anti-Ga-chelate Mab might be suitable for pretargeted immunoscintigraphy with the short-lived positron-emitter 68Ga.     

Development of an in vitro model for assessing the in vivo stability of lanthanide chelates

An in vitro model was developed to evaluate the in vivo stability of lanthanide polyaminocarboxylate complexes. The ligand-to-metal ratios for the chelates EDTA, CDTA, DTPA, MA-DTPA (monoamide-DTPA) and DOTA with the lanthanides lanthanum, samarium, and lutetium were optimized to achieve > or = 98% complexation yield for the resultant radiolanthanide complexes. The exchange of the radiolanthanides from their EDTA, CDTA, DTPA, MA-DTPA and DOTA complexes with Ca(2+) was determined by in vitro adsorption and in vitro column studies using hydroxyapatite (HA), an in vitro bone model. In vitro serum stability of these radiolanthanide complexes was used as an additional indicator of in vivo stability, although the mechanism of instability in serum will be different than with bone. The in vitro studies were consistent with the expected findings that the smallest lanthanide (Lu) formed the most stable complexes. In vivo studies were done to validate the in vitro model. Biodistribution studies in normal CF-1 mice showed that in vivo stability of the complex (i.e., the more lanthanide remaining in complex form) could be assessed by a combination of the urinary, bone and liver uptake. For example, biodistribution studies demonstrate that high urinary excretion correlated with complex stability, while high liver plus bone uptake correlated with complex instability. The urinary excretion of the EDTA complexes decreased from (177)Lu to (140)La indicating a loss in stability in the direction of (140)La, consistent with the in vitro studies. The more stable a lanthanide complex is, the lower its exchange with HA in vitro will be, and the lower its combined bone plus liver uptake and higher its urinary excretion will be in vivo. This investigation indicates that the in vivo stability can be determined by a screening method that measures the degree of exchange from the lanthanide chelate with hydroxyapatite (HA) and its serum stability.     

Lanthanide-based susceptibility contrast agents: Assessment of the magnetic properties

The T₂* contrast efficacy of paramagnetic contrast agents is dependent on their magnetic properties. Vibrating sample magnetometry (VSM) and the Live Chan NMR method have been used to evaluate the influence of ligand structure on the bulk magnetic susceptibility (BMS) of low-molecular weight (LMW) lanthanide chelates. VSM was also used for the BMS assessment of LMW lanthanide chelates covalentiy attached to cross-linked starch particles. The ligand structure had no influence on the BMS of the gadolinium (Gd) and dysprosium (Dy) chelates. The mean BMS value of the Dychelates was 1.8 fold higher than that of the Gd-chelates. The holmium (Ho) OTPA-BMA chelate had a similar BMS to that of Dy-DTPA-BMA while the lowest BMS was found for europium (Eu(lll)) DTPA-BMA. The covatent attachment of Gd-DTPA and Dy-DTPA to a cross-linked starch particle had no impact on their intrinsic magnetic properties. The BMS data were in good accordance with those obtained for non-particulate bound LMW Dy- and Gd-chelates. The magnetic susceptibility of the Gd-OTPA labeled particles was described by the Curie law, indicative of no magnetic interactions between Gd-DTPA molecules. The magnetic susceptibility of the Dy-DTPA labeled particles followed the Curie-Weiss law with a Curie-Weiss temperature of about -2 K, indicating magnetic interactions. The magnetic susceptibility of Dy-DTPA will, however, not be affected by such magnetic interactions at physiological temperatures.     

Improved route for the visualization of stem cells labeled with a Gd-/Eu-chelate as dual (MRI and fluorescence) agent.

A simple labeling procedure of stem/progenitor cells based on the use of Gd-HPDO3A and Eu-HPDO3A, respectively, is described. The Gd-chelate acts as T(1)-agent for MRI visualization, whereas the corresponding Eu-chelate acts as reporter in fluorescence microscopy. Owing to their substantial chemical equivalence, the two chelates are equally internalized in EPCs (endothelial progenitor cells), thus allowing their visualization by both techniques. The lanthanide chelates are entrapped in endosomic vesicles and the labeled cells retain biological activity with preservation of viability and pro-angiogenesis capacity. Hyperintense spots in MR have been observed for Gd-labeled EPCs injected under mice kidney capsule or grafted on a subcutaneous Matrigel plug up to 14 days after transplantation.     

Towards models for analysis of time-resolved luminescence spectra from quartz

Simple and stretched-exponential models for use in the analysis of time-resolved luminescence spectra from quartz are compared. The methods have been applied to evaluate lifetimes from portions of time-resolved luminescence spectra measured either during or after the luminescence stimulating pulse. We discuss the basis for use of the exponential functions as well as examine the application of, in this context, atypical stretched-exponential functions for analysis of time-resolved luminescence spectra from quartz.     

中国人镧系元素膳食摄入量和主要器官及组织负荷量研究

目的 测定我国成年男子膳食食品和主要器官、组织中镧系元素浓度和估算其膳食日摄入量和器官、组织负荷量。方法 在我国不同膳食类型4地区采集21例急死正常尸体10种主要器官、组织样品，连同过去所采当地食品和31例尸体6种器官、组织样品，采用ICP—MS或INAA法及必要的质量控制措施，分别测定了11和14种镧系元素浓度。按当地膳食组成和中同参考人器官组织重量估算了摄入量和器官、组织负荷量。结果 获得了我国12类食品和10种主要器官、组织样品中14种镧系元素浓度、摄入量和相应器官、组织负荷量。结论 本研究首次获得我国各类食品和成年男子10种器官组织中11种镧系元素浓度、相应膳食口摄入量和负荷量。所获结果为确定中国参考人镧系元素相应参数提供较前更为系统的依据，也为不同镧系元素、食品和器官组织提供了比较和背景值的国情资料。     

Diagnosis of militarily relevant diseases using oral fluid and saliva antibodies: fluorescence polarization immunoassay

This laboratory is developing fluorescence polarization (FP) methods as diagnostic tools to assay antibodies in saliva and other oral fluids. FP provides quantitation of molecular interaction, such as antigen-antibody binding, of a single, small-volume sample in real time and without prior separation of components such as blood cells. There is potential for wide-spread use of these homogeneous assays as noninvasive tests, especially as more compact, simplified fluorescence polarimeters become available. FP tests can be designed that are applicable to a wide spectrum of microorganisms and may be used in a clinic or far-forward deployed setting to aid in diagnosis of disease or verification of vaccination. Rapid salivary diagnostics, including FP, have been identified by the Office of Naval Research as requirements for future naval capabilities in basic and applied medical research for warfighter protection in casualty prevention. The applications of FP salivary diagnostics for determination of tuberculosis exposure and of anthrax immunization status are discussed as examples.     

Metabolism of indium chelates attached to monoclonal antibody: minimal transchelation of indium from benzyl-EDTA chelate in vivo

The metabolism of radiolabeled antibodies is important for radioimmunoimaging and therapy. The loss of indium-111 (111In) from the chelate can pose problems in imaging and increases the radiation dose to normal tissues. We have evaluated the loss in vivo of 111In from Lym-1-benzyl-EDTA-111In (an antibody conjugated with isothiocyanato-benzyl-EDTA) in normal mice. A monoclonal antibody (CHA 255) that binds to benzyl-EDTA-indium chelates, but not to other forms of indium, was used to measure the percent of 111In remaining in the chelate. Four days after injection, 97.4 +/- 2.2% of the 111In in the liver was still in the benzyl-EDTA chelate, as was 99.4 +/- 0.7% of the 111In in the urine, and 99.1 +/- 0.7% of the 111In in the blood. Studies in vitro indicate that a benzyl-EDTA-111In-antibody-chelate conjugate is more stable in human serum than a benzyl-DTPA-111In conjugate, and that both benzyl-chelate conjugates are much more stable than an unsubstituted DTPA conjugate.