In situ observation of the dissolution kinetics of Al_(2)O_(3) particles in CaO–Al_(2)O_(3)–SiO_(2) slags using laser confocal scanning microscopy

The dissolution kinetics of Al_(2)O_(3) in CaO-Al_(2)O_(3) SiOslags was studied using a high-temperature confocal scanning laser microscope at 1773 to 1873 K.The results show that the controlling step during the Al_(2)O_(3) dissolution was the diffusionin molten slag.It was found that the dissolution curves of Al_(2)O_(3) particles were hardly agreed with the traditional boundary layer diffusion model with the increase of the CaO/Al_(2)O_(3) ratio of slag.A modified diffusion equation considering slag viscosity was developed to study the dissolution mechanism of Al_(2)O_(3) in slag.Diffusion coefficients of Al_(2)O_(3) in slag were calculated as 2.8×10to 4.1×10m~2/s at the temperature of 1773-1873 K.The dissolution rate of Al_(2)O_(3) increased with higher temperature,CaO/Al_(2)O_(3),and particle size.A new model was shown to be v_(Al_(2)O_(3))=0.16×r_(0)^(1.58)×x^(3.52)×(T-T_(mp))^(1.11)to predict the dissolution rate and the total dissolution time of Al_(2)O_(3) inclusions with various sizes,where vAl_(2)O_(3) is the dissolution rate of Al_(2)O_(3) in volume,μm^(3)/s;x is the value of CaO/Al_(2)O_(3) mass ratio;R_(0) is the initial radius of Al_(2)O_(3),μm;T is the temperature,K;T_(mp) is the melting point of slag,K.

Confocal Laser Scanning Microscope Evaluation of Early Bacterial Colonization on Zirconium Oxide and Titanium Surfaces:An in vivo Study

To investigate the bacterial colonization on zirconium oxide and titanium surfaces in vivo quantitatively using a confocal laser scanning microscope （CLSM）. Ten samples of zirconium oxide ceramic and commercially pure titanium were fabricated and polished using silicon carbide abrasive paper. One sample from each group was evaluated topographic pattern under a scanning electron microscope. One sample from each group was to evaluate roughness using a profilometer. Eight volunteers were selected. The samples were cemented at the buccal surfaces of upper first molars. All samples were removed after 48 hours, immersed in SYTO-9 and propidium iodide fluorescent to stain for adherent bacteria and obseIved with CLSM. Fewer bacteria were observed in zirconia group than titanium group. However, there was no statistical difference between two groups. The experimental results demonstrate that zirconium oxide may be considered as a promising material for dental implant abutments.

Quantum Dots as Fluorescent Labels for Detection of Heat Shock Protein in Tumor Tissue Using Laser Scanning Confocal Microscope

A new Quantum Dots（Qdots） nanocrystal composed of semiconductor core and zinc sulfide shell, and its feasibility as labels in immunofluorescence analysis for the imaging of tumor biomarkers by laser scanning confocal microscope（LSCM） was investigated. Qdots taged by mercaptoacetic acid were conjugated with second antibody, then imaging differences of Heat Shock Proteins 70（HSP70） in renal carcinoma tissure sections with immunofluorescence analysis method using Qdots bioconjugates and conventional organic dye FITC were observed by LSCM to assess the brightness and opticalstability of Qdots. The experimental results showed Qdots bioconjugates achieved the better results in demonstrating HSP70 with more brighter color and more clear picture than FITC labels. Moreover, the label signals of Qdots did not fade clearly after continued exposure to a 488 nm laser for 1 h. The Qdots bioconjugates have good feasibility in immunofluorescence analysis for the bioimaging by LSCM.

Qualitative analysis of re mineralized carious lesions subjected to fluoride supplement through confocal laser scanning microscope

Aim: 1] Comparative evaluation of the linear depth of induced remineralized lesions after subjecting to fluoride supplements and 2] To assess the average fluorescence at both the demineralized and the remi-neralized zones in all the three study groups under confocal laser scanning microscope. Method: Forty five sound human premolars extracted for orthodon-tic reasons were decoronated 1 mm below the ce-mento-enamel junction and coated with nail varnish except for a 3 × 3 mm window on the buccal surface. The samples were placed in 50 ml of de mineralizing solution at pH 4.6 for 96 hours. Following deminera-lization, the lower half of the 3 × 3 mm window in all the samples were covered with nail varnish to serve as control. The samples were randomly divided into three groups of fifteen teeth each (n = 15) and speci-mens in group A[Nfd] were remineralized using non-fluoridated dentifrice [control], those in groups B [Fd5] and group C [Fd10] using 500 ppm and 1000 ppm of fluoride containing dentifrice, respectively. The specimens were subjected to a 20 day reminera-lization treatment regimen and were sectioned into 100 μm thick sections and two images were captured on the buccal surface from either side of the midpoint of occluso-cervical length using confocal laser scan-ning microscope [CLSM]. Results: were tabulated and statistically analyzed by Anova. Study concluded that 1000 ppm fluoridated dentifrice showed a greater degree of remineralization than other groups and confocal laser scanning microscopes gives promising results in the diagnosis of early enamel lesions over the conventional methods.

Highly specific characterization and discrimination of monosodium urate crystals in gouty arthritis based on aggregation-induced emission luminogens

Existing technologies used to detect monosodium urate(MSU)crystals for gout diagnosis are not ideal due to their low sensitivity and complexity of operation.The purpose of this study was to explore whether aggregation-induced emission luminogens(AIEgens)can be used for highly specific imaging of MSU crystals to assist in the diagnosis of gout.First,we developed a series of luminogens(i.e.,tetraphenyl ethylene(TPE)-NH_(2),TPE-2NH_(2),TPE-4NH_(2),TPE-COOH,TPE-2COOH,TPE-4COOH,and TPE-Ketoalkyne),each of which was then evenly mixed with MSU crystals.Next,optimal fluorescence imaging of each of the luminogens was characterized by a confocal laser scanning microscope(CLSM).This approach was used for imaging standard samples of MSU,hydroxyapatite(HAP)crystals,and mixed samples with 1:1 mass ratio of MSU/HAP.We also imaged samples from mouse models of acute gouty arthritis,HAP deposition disease,and comorbidities of interest.Subsequently,CLSM imaging results were compared with those of compensated polarized light microscopy,and we assessed the biosafety of TPE-Ketoalkyne in the RAW264.7 cell line.Finally,CLSM time series and three-dimensional imaging were performed on MSU crystal samples from human gouty synovial fluid and tophi.As a promising candidate for MSU crystal labeling,TPE-Ketoalkyne was found to detect MSU crystals accurately and rapidly in standard samples,animal samples,and human samples,and could precisely distinguish gout from HAP deposition disease.This work demonstrates that TPE-Ketoalkyne is suitable for highly specific and timely imaging of MSU crystals in gouty arthritis and may facilitate future research on MSU crystal-related diseases.

Dispersion of particles in the coatings characterized by laser scanning confocal micrscopy(LSCM) I:Vertical dispersion of particles in the coatings and the weathering property studied by orthogonal analysis method of LSCM

Two kinds of TiO_2 filled epoxy coatings were designed and prepared to obtain pigments with different dispersion degrees of TiO_2 particles.Laser scanning confocal microscope(LSCM)was used to investigate both the horizontal and vertical distributions of TiO_2 particles in the coatings.The results indicated that TiO_2 in the two samples shared considerable similarity in horizental dispersion,but exhibited great difference in vertical dispersion.TiO_2 showed uniform vertical distribution in disp coating,wheras a gap about 1.1μm was found in the non-disp coating,which significantly influenced the surface optical properties of the coatings during weathering.Based on the confocal data,the model of dispersion of pigments in the coatings was proposed and the change of surface properties during weathering was predicted:the surface optical properties showed an initial decrease followed by a subsequent increase,which was in good agreement with the weathering data.

Using laser confocal scanning microscope to study ischemia-hypoxia injury in rat brain slice

The level of lipid peroxidation and cellular necrosis in rat living brain slices during brain ischemia-hypoxia injury have been observed using a laser confocal scanning microscope (LCSM) with double labeling of fluorescent probes D-399 (2, 7-dichlorofluorescin diacetate) and propidium iodide (Pl). The hypoxia and/or reoxygenation injury in rat brain slices is markedly decreased by pretreatment with L-NG-nitro-arginine (L-NNA) and N-acetylcysteine (NAC), showing that the nitric oxide (NO) and other free radicals play an important role in brain ischemia-hypoxia injury.

成年鼠缺血性脑损伤诱导nestin的表达

应用免疫组化和免疫荧光双标技术结合激光共聚焦扫描显微镜,观察缺血性脑损伤后脑内nestin的表达及其细胞类型。实验观察结果为,再灌后1天,在缺血中心区可见nestin阳性突起;再灌后3天和1周时,除缺血中心区外,周边Ⅰ、Ⅱ、Ⅲ区均有nestin大量表达;而2周时仅限于周边Ⅰ区大量表达。激光共聚焦扫描显微镜观察到,再灌后3天,周边Ⅰ区 nestin阳性突起主要与GFAP共存;2周时,nestin阳性突起变粗、变长,并与NSE的共存明显增多。上述研究结果提示,脑缺血可诱导大鼠脑缺血区域表达nestin,该表达可能与神经细胞的修复有关。

15-HETE对肺动脉平滑肌细胞钙离子浓度的影响

目的 通过阻断细胞外钙离子 (Ca2+ )内流,观察 15 羟化二十烷四烯酸 ( 15 HETE)对兔肺动脉环收缩张力和肺动脉平滑肌细胞内游离钙浓度 ( [Ca2+ ]i)的影响,探讨缺氧时 15 HETE引起细胞钙动员的机制。方法 采用组织浴槽血管环方法观察L 型钙通道阻断剂硝苯地平和无钙液对15 HETE诱导的兔肺动脉环收缩力的影响;酶法分离培养兔肺动脉平滑肌细胞,激光扫描共聚焦显微镜测定 15 HETE对细胞内 [Ca2+ ]i的作用。结果 在正常组和缺氧组, 10μmol·L-1硝苯地平和无钙液对 1μmol·L-1 15 HETE引起的肺动脉环收缩均没有影响;培养的 15 HETE组细胞 (正常培养的细胞暴露于 1 μmol·L-1 15 HETE下继续孵育 8min)与正常对照组相比,细胞内 [Ca2+ ]i明显增加 (P<0 05)。结论 15 HETE可引起肺动脉平滑肌细胞 [Ca2+ ]i增加,且此钙来源于细胞内贮钙库的释放。

MAP30苦瓜抗HIV蛋白对细胞内HBeAg表达抑制作用的定量分析

目的 :研究Mr 为 30 0 0 0的苦瓜抗HIV蛋白 (MAP30 )对 2 .2 .15细胞内HBV基因表达的抑制作用。方法 :将HBVDNA转染的人肝癌细胞株 2 .2 .15 ,在MAP30存在的情况下培养后 ,进行间接免疫荧光染色 ,用共聚焦技术定量分析细胞内HBeAg的表达。结果 :与对照组相比较 ,MAP30培养的细胞内HBeAg荧光量明显减少。结论 :MAP30对 2 .2 .15细胞内HBeAg的表达有明显的抑制作用。

用共聚焦激光扫描显微镜观测GFP标记的内生固氮菌Klebsiella oxytoca SA2侵染水稻根

用高效绿色荧光蛋白 (greenfluorescentprotein ,GFP)突变体EGFPmut2的基因标记内生固氮菌———产酸克雷伯氏菌 (Klebsiellaoxytoca (Fl櫣ｇｇｅ)Lautrop)SA2 ,用标记菌接种限菌培养条件下生长的水稻 (OryzasativaL .)幼苗 ,在接种后 1、2、4、8、12、16和 2 1d ,用共聚焦激光扫描显微镜对水稻鲜根进行光学切片 ,显示了标记菌从水稻根成熟区表面向根内入侵的过程。定殖在根表面的标记菌主要从侧根伸出的位置侵入侧根皮层 ,从邻近发生侧根的位置进入内皮层和维管柱。SA2还能从初生根成熟区无侧根伸出的位置侵入皮层向维管柱迁移。SA2入侵水稻根引发了水稻根局部的过敏反应以阻滞细菌进一步入侵 ,表现为侵入根内的细菌周围的根细胞细胞壁变厚 ,在蓝光下发出很强的黄绿荧光。

香豆素类成分在球花石斛(Dendrobium thyrsiflorum Rchb.f.)根中的分布和相对含量

利用激光共聚焦显微技术(laserscanningconfocalmicroscopy,LSCM)观测了香豆素类成分(coumarins)在球花石斛(DendrobiumthyrsiflorumRchb.f.)根中的分布及相对含量,为合理利用药用石斛提供科学的理论依据。观察结果显示香豆素类成分主要分布于球花石斛根的细胞壁,尤以细胞的角隅处为多;自发荧光强度以外皮层的最高,根被的最低。

激光扫描共聚焦显微镜(LSCM)在植物细胞学中的应用

激光扫描共聚焦显微镜(LSCM)有效地排除了非焦平面信息,提高了分辨率及对比度,使图像更为精确清晰。目前已广泛应用于植物细胞学研究,在植物药研究方面有着广阔的应用前景。综述近年来的相关研究以反映其最新进展。

中药HB对喉癌Hep-2细胞的抑制作用及其机制研究

目的 研究蜈蚣提取物HB对喉癌Hep 2细胞生长的抑制作用及其作用机制。方法 MTT法观察不同浓度的HB对Hep 2细胞的生长抑制情况 ;采用激光扫描共聚焦技术分析HB作用Hep 2细胞 48h后细胞内钙、DNA含量的变化情况。结果 Hep 2细胞用 10 μg/ml、10 0 μg/ml的HB处理 72h后 ,细胞存活率显著降低 ;用 10 μg/ml和2 0 μg/ml的HB处理Hep 2细胞 48h后 ,细胞内钙含量显著升高 (P <0 .0 0 1) ,DNA含量显著降低 (P <0 .0 0 1)。 结论 中药HB在体外对Hep 2细胞的生长有明显地抑制作用 ,机制与降低细胞内DNA含量 。

采用Fluo-3 AM-ester探针研究桃不同组织的Ca^2＋信号分布

钙作为植物最重要的矿质元素之一,在植物生理和生化活动中起重要作用。采用激光扫描共聚焦显微镜,以Fluo-3 AM-ester为荧光探针研究了桃果肉和叶柄组织中Ca2+的分布,结果表明Fluo-3 AM-ester可作为荧光探针对桃不同组织中游离的Ca2+信号强度和分布特征进行分析。不同的物镜观察(10×和20×)下,Ca2+荧光信号相对强度差异不大;在叶柄中Ca2+荧光信号主要分布于维管束,在果实中多分布于细胞壁;但叶柄中荧光信号分布明显高于果肉组织,这与钙在不同组织中的功能有关。

荧光探针Fluo-3/AM和FuraRed测量单细胞中比率钙

应用ACAS570激光扫描共聚焦显微镜（laserscanningconfocalmicroscope，LSCM简称共焦显微镜）和钙离子指示剂Fluo－3／AM、FuraRed荧光探剂双标记技术，测定了单个活细胞内比率钙的动态变化。结果显示37℃，Fluo－3／AM浓度10μmol／L，FuraRed浓度10μmol／L的条件下，昆明小鼠巨噬细胞负载1h左右即可获良好的标记效果。用比率探针测量细胞内Ca^2＋的动态变化，使Ca^2＋的定性定量测定不受染料浓度、细胞大小、照射光强度以及一定程度的光漂白和染料泄漏等因素的影响，提供了一种准确定性定量细胞内Ca^2＋的实时、动态、原位变化的新方法。

成年大鼠液压冲击脑损伤诱导室下区Nestin和GFAP的表达

目的 应用免疫组织化学和免疫荧光双标技术结合激光共聚焦显微镜 ,观察脑损伤后不同时期室下区巢蛋白(Nestin)和胶质酸性纤维蛋白 (GFAP)的表达及其细胞类型。方法 用动液压冲击法建立动物模型 ,用LeicaQ5 0 0IW图像处理系统及免疫组化和免疫荧光双标法分析和显示不同时期 (1、3、5d ,1、2、3、4、6、8周 )Nestin和GFAP的表达变化。结果 损伤 2 4h后 ,室下区有大量Nestin和GFAP的表达 ,且激光共聚焦显微镜观察到有Nestin阳性突起与GFAP共存现象 ,7d左右达到高峰 ,然后随着时间的推移其表达逐渐下降。正常组和假手术组 ,室下区也可见到Nestin和GFAP的表达 ,但与脑损伤组相比较低 (P <0 .0 5 )。结论 脑损伤可诱导室下区Nestin和GFAP的表达 ,Nestin阳性突起与GFAP共存细胞可能具有干细胞特性 ,与神经元的修复有关。

家蝇天蚕素对人肝癌BEL-7402细胞的作用靶点

将家蝇天蚕素(终浓度50μmol/L)加入对数生长期的人肝癌BEL-7402细胞中作用12h,扫描电镜观察天蚕素对人肝癌细胞细胞膜的影响。在此基础上,采用绿色荧光素异硫氰酸(FITC)标记天蚕素,运用激光共聚焦显微镜观察FITC标记的天蚕素与人肝癌细胞细胞膜的作用。肿瘤细胞经天蚕素作用12h后,扫描电镜显示细胞膜表面微绒毛大部分消失;激光共聚焦显微镜显示大部分天蚕素与细胞膜结合,细胞内也发现部分天蚕素进入。

LSCM观察三七总皂甙及其组分对游离钙离子影像变化的影响

用盖玻片或ＭａｔＴｅｋ培养皿培养猪主动脉内皮细胞（ａＥＣ）约１８ｈ后，以荧光探针（ｆｌｕｏ－３／ＡＭ终浓度１０μｍｏｌ·Ｌ－１）负载，当细胞内游离钙离子（Ｃａ２＋）与ｆｌｕｏ－３结合后，通过激光扫描共聚焦显微镜（ＩｎＳＩＧＨＴ－ｐｌｕｓＩＱ型）观察细胞的荧光影像变化。基于三七总皂甙（ｔ－ＰＮＳ）对ａＥＣ所产生的生物活性物质有明显的作用，本拟进一步探索上述药物对ａＥＣ中［Ｃａ２＋］；的影响。实验结果显示：在ＡＴＰ作用后，ｔ－ＰＮＳ可使ａＥＣ内的荧光强度在２０～８０ｓ内急剧升高至峰值。但其组分Ｒｂ１及Ｒｇ１使荧光强度有下降或无明显变化。提示ｔ－ＰＮＳ有使内皮细胞［Ｃａ２＋］；升高的作用。

细胞内微丝骨架的变化对人肝癌Bel-7402细胞形态的影响

癌细胞的形态及癌细胞的生物学行为(如粘附、迁移和浸润)与细胞内骨架系统密切相关,本研究利用Phalloidin FITC标记人肝癌Bel 7402细胞内F 肌动蛋白(F actin),使用激光扫描共聚焦显微镜观测细胞微丝骨架与癌细胞形态的关系,及Cyt B处理Bel 7402细胞后,癌细胞内微丝骨架的变化。结果显示:癌细胞内F actin解聚,F actin小体的聚集重组是癌细胞的形态变化的主要因素之一。