电镜超薄切片批量染色的新方法

超薄切片染色是透射电镜生物样本制备中的关键环节。本文介绍了一种透射电镜超薄切片染色的新方法,利用简单自制的染色液防污存储与批量染色装置,可对超薄切片载网进行批量化染色,不但极大提高染色效率,而且减少了染色过程中染色液、样品的污染,达到染色液的重复利用,同时减少重金属对操作者的伤害和对环境的污染。     

基于Ansys/Flotran的夹持型超薄切片染色板优化设计

电镜超薄切片样品制备中,重金属染色是不可缺少的关键步骤.本文基于夹持型超薄切片染色板工作原理,建立4种不同染色孔形状的染色板模型(染色孔底部与侧壁夹角90°,105°,120°和135°),对水流冲洗染色板过程进行二维模拟,采用Ansys/Flotran方法计算超薄切片表面的实际冲洗液流速度和湍流动能.结果表明,随着染色孔底部与侧壁夹角的增加,超薄切片表面的水流冲洗速度逐渐增加,且不同染色孔内的湍流动能逐渐变得均匀,在染色孔的壁与底面的夹角为105°时,湍流动能最大,即水流与超薄切片表面残余的重金属染液之间交换程度最大,此结构被确定为最优化设计.根据计算流体动力学研制的产品,操作简便、安全有效,为提高透射电镜的样品染色效率和质量提供理论基础.     

电镜超薄切片真空染色法

介绍一种简便易行的超薄切片真空染色法。使用简单易得的工具及自制装置-真空保鲜盒、染色硅胶板和染色硅胶板架,能够得到较好的染色效果。结果显示,与目前现有的传统方法相比较,超薄切片真空染色法省时、省试剂,染色效果好,一次可进行大批量切片染色(一次可达200张超薄切片)。     

一种简易的超薄切片染色器

本文介绍了一种简易的超薄切片染色器的制作和使用方法。各实验室都有制作比染色器所需的材料,其制作和使用方法简单。可进行铀、铅单、双重染色。染色效果良好。在透射电镜生物样品超薄切片染色(尤其铅染)过程中,往往由于CO_2的影响,常在超薄切片上产生碳酸铅沉淀,使切片受到污染,影响观察,严重时无法观察使之前功尽弃。我们参考LKB2160型染色机的工作原理和染色过程,结合工作中的实践,自制了一种简易的染色器,可进行铀、铅单、双重染色。使用此染色器,可使超薄切片的染色达到防污染、规格化、省时间的效果。现将其制作及使用方法介绍如下:     

高分子材料冷冻切片的捞片技术

长期以来,在透射电镜样品制备超薄切片中,常规的捞片方法是采用带支持膜的载网收集。近年来,随着电镜的广泛应用,冷冻切片技术在高分子材料中应用也越来越广泛,由于冷冻切片技术要求较高,难度也大,切片厚度的控制是关键,否则在电镜下电子束将无法穿透过去。因此,在切片过程中,除了掌握好制样重要技术环节以外,要求尽可能采用无膜载网进行捞片,以减少电子束穿透厚度和切片的热漂移,这样才能保证获得厚度低于100nm的切片。本人经过长期的工作实践不断的摸索和总结,现将冷冻切片技术中应用无膜载网收集切片应掌握的技术介绍如下。冷冻切片样品…     

透射电镜电子染色方法的改良

为提高透射电镜生物样品超薄切片的染色效率,在传统染色方法基础上进行改良,使用自制染色装置,在较短时间内完成大批生物样品超薄切片的电子染色。与传统方法相比较省时省药,减少污染机率,电镜观察的超微结构清晰,反差较好。     

一种高效的超薄切片染色法

本文介绍使用“连通式”玻管染色装置(如图)的超薄切片染色法及在染色中应注意的事项。经多次使用证明,用该法染出的超薄切片电子着色均匀,结构清晰,有良好的反差。该法能保证切片染色条件的一致,有效地防止切片染色污染,提高染色质量。用该法染色,染液用量省,染20张载网只需1ml染液,降低了铀、铅污染环境的可能性。染色和冲洗时间大大缩短,染30张载网前后共只需30分钟,大大提高了工作效率。该染色装置由三部分组成,制作如下:A.染色管取一根长130mm、内径6mm的玻管,用酒精喷灯加热玻管的一端至内径缩为1mm。在距另一端20mm处,用喷射火焰打一个     

一种用硅橡胶载网夹进行超薄切片染色的有效方法

本文介绍了一种能减少污染、省时省力的电镜超薄切片染色方法。     

提高电镜超薄切片染色效率的自制装置

用传统的方法进行透射电镜超薄切片染色费时费工,且易造成污染,本文介绍一种自制的染色装置能很好地解决这一问题,提高染色效率,不同生物样品的超微结构保存良好、清晰.     

立体高压电子显微术的生物学应用

超薄切片法的局限性在于它所形成的图象只有二维性质。运用高压电镜使得观察的切片厚度大大增加成为可能。然而,切片越厚,在同一部位图象中重叠的结构也就越多,也就越难区别。选择性染色方法解决了这个问题,它对需要观察的感兴趣结构染色而其他结构相对地不染色。本文介绍了应用立体高压电镜术研究骨骼肌纤维的横管系统。     

耳蜗透射电镜超薄切片样品的制备技术改进

针对耳蜗组织透射电镜样品制备难度大,不易获得质量较好的超薄切片的问题,作者通过延长锇酸后固定时间和增加树脂渗透的浓度,改善了树脂包埋组织的切割性能.电镜图片显示,明显地提高了超微切片样品制备质量.     

A TEM study on melt-crystallized poly(butylene terephthalate)

A study was made to investigate an optimum condition for ruthenium tetraoxide staining of poly(butylene terephthalate) (PBT) with glass transition temperature as low as ca. 23 degrees C for TEM observation of melt-grown PBT crystals. The morphological study of ultrathin sections prepared based on optimum staining at 25 degrees C for 8 h revealed formation of fringed-micellar crystal nuclei in the early stage and folded-chain fringed-micellar crystals in the later stage of isothermal crystallization at 40 degrees C from melt. The fraction of chain-folded crystals increased with increasing crystallization temperature from the quantitative analysis of crystal thickness in comparison with the reported morphology using the replica method.     

Measurement of the repeat period of myelin sheath using ultrathin frozen sections

The myelin sheath of peripheral nerves was observed by transmission electron microscopy (TEM) using plastic-embedded sections and ultrathin frozen sections. Repeat distances of myelin sheaths were measured in high-powered electron micrographs. The ultrathin frozen sections showed a longer repeat distance than the plastic-embedded sections. The ultrathin frozen sections were thought to contain fewer artefacts, as they had not been subject to dehydration and embedding. It is known that broken myelin sheaths are often observed under conventional TEM. It is thought that these procedures cause contraction and partial destruction of the myelin sheath.     

煤岩样品TEM超薄切片制备方法研究

本文作者以泥炭、褐煤、烟煤及无烟煤为例,探讨了超薄切片制备方法对不同煤阶样品的适用性。在采用包埋切片法制备煤岩样品TEM超薄切片时,首先通过浸解离析方法和HCl、HF逐级化学浸蚀方法使煤中自然共生组合的有机显微组分离析及脱除无机矿物质;利用光学显微镜(透射/反射模式)镜检离析显微组分后,使用SPI812树脂对挑出的单一显微组分块体渗透及包埋聚合。显微组分块体形态学及嵌布矿物成分分析使用扫描电子显微镜的低真空二次电子像模式和EDS面分布分析模式;利用透射电子显微镜检查超薄切片效果。实验表明采用上述方法制备煤岩TEM超薄切片样品的成功率较高,并且能够比较真实的再现显微组分的微观结构。     

透射电镜神经组织制样包埋剂比较

为探讨两种常用包埋剂Epon 812和Eponate 12对神经组织透射电镜样品的影响,本文采用多聚甲醛-戊二醛溶液对wistar大鼠进行全身灌流固定后,取材海马、脊髓及坐骨神经,按常规透射电镜样品技术制作超薄切片.电镜观察显示,经Eponate 12包埋后的样品,在切片的透明度及超微结构的清晰度上均优于Epon 812包埋的超薄切片样品.由于Eponate 12的浸透效果好,有效地避免了有髓神经纤维髓鞘上出现孔洞结构的制样问题.本实验小组的实验结果表明:对于神经组织,尤其是周围神经组织的超薄切片样品,使用Eponate 12作为包埋剂,效果会更好一些.     

Correlative light microscopy, scanning electron microscopy, and transmission electron microscopy of osmium-macerated biological tissues

A method facilitating correlation of light microscopic (LM), scanning electron microscopic (SEM) and transmission electron microscopic (TEM) images was developed. Rat kidney and heart were initially subjected to the osmium maceration procedure and then embedded in acrylic resin. Semithin sections of the tissue blocks were first provided for LM and then examined by SEM after resin removal. Furthermore, the ultrathin sections adjacent to the semithin sections were observed by TEM. The three-dimensional images of intracellular organelles provided an informative adjunct to LM and TEM.     

Quantitative TEM characterizations of La/B4C and Mo/B4C ultrathin multilayer gratings by the geometric phase method

Artificially fabricated multilayers that exploit the effect of Bragg diffraction at long wavelengths are used as X-ray optical components. Periodic ultrathin bilayer stacks of, alternatingly, a metallic reflection layer and a non-metallic spacer layer prepared as μm-scaled surface grating promise particularly high performance in applications as X-ray filters for high-resolution spectroscopy. Such gratings can be prepared by multilayer deposition onto Si(0 0 1) gratings or by coating flat Si(0 1 1) substrate surfaces with a multilayer, followed by subsequent etching of a grating structure. The structural quality of ultrathin multilayer gratings of both types has been characterized quantitatively combining TEM bright-field imaging of specimen cross-sections and applying a geometric phase method. The geometric phase method has been originally developed for the analysis of local displacement fields from HREM images and allows to obtain the relevant structure parameters. The application of this method to the characterization of Mo/B₄C and La/B₄C multilayer systems shows that the functionally decisive structure parameters, such as layer perfection, layer periodicity, and layer orientation, can be obtained with high precision from evaluation of TEM bright-field images. The essential role of such data analyses for a quantitative characterization of multilayer systems and for optimising the layer deposition techniques in the fabrication of X-ray optical layer systems will be demonstrated and discussed.     

Direct observation of a stacking fault in Si(1 - x)Ge(x) semiconductors by spherical aberration-corrected TEM and conventional ADF-STEM

Spherical aberration (C(S))-corrected transmission electron microscopy (TEM) and annular dark-field scanning TEM (ADF-STEM) are applied to high-resolution observation of stacking faults in Si(1 - x)Ge(x) alloy films prepared on a Si(100) buffer layer by the chemical vapor deposition method. Both of the images clarify the individual nature of stacking faults from their directly interpretable image contrast and also by using image simulation in the case of the C(S)-corrected TEM. Positions of the atomic columns obtained in the ADF-STEM images almost agree with a projection of the theoretical model studied by Chou et al. (Phys. Rev. B 32(1985): 7979). Comparison between the C(S)-corrected TEM and ADF-STEM images shows that their resolution is at a similar level, but directly interpretable image contrast is obtained in ultrathin samples for C(S)-corrected TEM and in slightly thicker samples for ADF-STEM.