Cryostat Sectioning; Modification of the Antiroll Plate

A replaceable antiroll plate and holder have been designed for use in the Ames Lab-Tek cryostat which replace the plastic plate supplied with the instrument and insure a flawless, properly aligned plate for maximum efficiency in thin section cutting. A metal plate holder is attached to the existing screw-driven bracket provided with the instrument by the manufacturer. Glass plates made from one half of a 1.5 × 3 inch microscope slide are coated on the leading edge with spray-on Teflon and provided with tape spacers. These plates slip into the holder and can be adjusted for angular inclination and alignment with the cutting edge by movement within the holder or manipulation of the adjustment screw.     

Capsules for Frozen Sectioning of Small Specimens

It was necessary to make sections of small unfixed specimens which had been frozen while still immersed in their normal culture medium. The principal difficulty stemmed from the poor sectioning quality of the frozen culture medium. A capsule is described which has a narrow well in which the tissue specimen fits snugly within a small amount of culture medium. After freezing, the whole capsule is sectioned and the resulting sections, being nearly devoid of culture medium, are of good quality.     

介绍一种半薄切片定位样品的方法

介绍了一种半薄切片定位样品的方法。此法与前人的方法不同之处在于位于载玻片之上被重新包埋于塑料环内的切片与载玻片的分离真人处步骤是“切片被包埋聚合好后,立即从６０￣６５℃温箱内被转入冰箱冷冻室中（－１８℃）放置５￣１０分钟。然后,将载玻征从冷冻室中取出,轻推塑料环,即可使包埋在塑料环内的切片与载玻片分离。这一方法成功地解决了样品中靶细胞的发育时期确定和样品丢失问题。而且还有简单、易操作和成功率高等优     

Drinking-Straw Molds for Embedding Multiples of Longitudinal Segments for Cryostat Sectioning

It is often desirable to obtain cryostat cross sections of tubular structures, such as oviduct, uterus, esophagus, or small intestine, and to retain the sequential relationships of such sections. In this laboratory the following technic which has been used on rat and mouse tissues from newborn and weanling animals has proved to be rapid, inexpensive, and productive of uniformly excellent results.     

Embedding Thin Plant Specimens for Oriented Sectioning

Small plant structures such as small primary roots, filamentous mosses and algae are difficult to orient for sectioning since they become wavy and curl during embedding. A method is described for embedding and orienting tiny plant specimens in a glycol methacrylate resin using self-constructed flat molds. Prior to sectioning, small samples can be oriented in both the longitudinal and the transverse plane. As several samples can be sectioned simultaneously, time-consuming trimming of the blocks is reduced substantially. The efficiency of this technique has been demonstrated using the tiny roots of the model plant Arabidopsis thaliana (L.) Heynh.     

Cryostat Techniques: Methods for Improving Conservation and Sectioning of Tissue

Tissues have been conserved for satisfactory enzymatic histochemical assay for up to 12 mo by low temperature storage (in a dry ice chest or at -40° C) and by measures designed to offset the deleterious effects of sublimation of H₂O by using the following modifications of standard procedures (which include steps to give a mechanical support to otherwise fragmenting sections): 1. tissue blocks are coated with a polystyrene solution between each storage period; 2. modified bolts are used as tissue holders and types of bolt holders have been designed to fit on standard microtomes which permit manipulation of each tissue block independently of its mates on the same bolt holder, or the simultaneous cutting of all blocks on any one holder with each advance of the microtome feed; 3. tissues are coated with 20% polystyrene in methylene chloride prior to cutting and rubber cement painted on the slide as an adhesive. Lillie's 20% polystyrene diethylbenzene is used as a mounting medium. Other details of practical importance include the technique of freezing, control of moisture within a cryostat and on the microtome, and tests on histochemical procedures.     

Estimation of Thy-1 in Cryostat Sections of Nervous Tissue

The conventional assay for measuring cell surface antigens--the quantitative absorption of antibody by tissue homogenates--proved inadequate when used to determine the level of Thy-1 glycoprotein in rat nerves and peripheral ganglia. In this paper we report that the binding of 125I-labelled Fab fragments of a monoclonal anti-Thy-1 antibody to cryostat sections is sufficiently sensitive to give consistent estimates of the Thy-1 level on single samples of even small nerves. Observed levels of Thy-1 were generally higher than had previously been thought, and in particular we found no nerves totally lacking the antigen. The lowest levels (6-10 pmol/mg protein) were in peripheral nerves with a large motor component. Autonomic and sensory nerves had higher levels (15-20 pmol/mg protein). The highest levels were on the optic nerve (34 pmol/mg protein), superior cervical sympathetic ganglion (40 pmol/mg protein), and the cerebellar vermis (46 pmoles/mg protein; the only brain region examined in this study). From a practical point of view, the cryostat assay has the advantage that measurements of Thy-1 can be done on sections from the same series as is used for immunohistochemical localisation.     

A Method for Thin Sectioning Heavily Sclerotized Cuticle of Odontotaenius disjunctus

Elytra of the bessbeetle, Odontotaenius disjunctus were thin sectioned after embedding in epoxy resin. Sections were cut with a diamond saw, ground to the desired thickness on a rotary grinder and polished. Tearing and distortion were reduced when compared to knife-cut sections of heavily sclerotized cuticle.     

Macroscopic Sectioning of Undecalcified Tissues

An inexpensive laboratory apparatus designed to section undecalcified teeth and bone has been constructed. It consists of an electric motor with a mandrel bearing a carborundum sectioning disk centered within a Plexiglas enclosure. A coolant flows from a reservoir positioned in the upper portion of the Plexiglas enclosure to prevent desiccation or burning of the tissues. Undecalcified tissue can be cut into a series of thin, 0.5-1 mm slices to facilitate studies of pulpal enzymes, tooth morphology and design of dental cavity preparations.     

高山榕染色体制片优化及核型分析

以高山榕种子的根尖为材料进行染色体常规压片,比较不同预处理方法和解离方法对高山榕染色体制片的影响,以选择最优的压片方法制片并进行核型分析。结果显示,预处理24 h的总体效果优于12 h;3种预处理液的总体作用效果为0.05%秋水仙素>混合液>0.002 mol.L-18-羟基喹啉,综合比较后认为混合液低温4℃处理24 h为最佳预处理方法。解离方法选择1 mol.L-1HCl中60℃水浴2~3 min较为适宜。首次报道了高山榕核型公式为2n=2x=30=22m(2SAT)+6sm+2T,核型不对称系数为61.54%,核型属于Stebbins核型分类中的2C类型。     

植物冰冻切片条件的优化及其与石蜡切片在组织化学应用中的比较

冰冻切片是植物组织学研究中一项重要的实验技术,冷冻温度和冷冻时间是决定切片质量的关键因素。通过比较15种冰冻切片条件,得出植物组织直接冰冻切片较适宜的冷箱温度、冷台温度和冷冻时间。同时,通过对5种植物的不同组织进行组织化学染色,比较了新鲜材料直接冰冻切片与常规石蜡切片在不同化学成分鉴定上的异同及各自的适用范围。结果表明,对于多糖、蛋白质和角质,两种切片方法的鉴定结果比较一致,但对于脂肪只能采用冰冻切片技术。研究结果对植物组织学实验和研究方法的改进具有一定的参考价值。     

应用Steedman's wax切片法观察植物细胞微管骨架

微管骨架在植物发育过程中起重要作用.由于植物细胞的特殊性,与动物细胞相比植物微管骨架的研究遇到更多的困难.简略地介绍了曾被国内外学者应用的植物微管骨架的各种研究方法及其局限性.Steedman's wax是一种多脂蜡.它熔点低(35～37℃),具有与石蜡相同的切片性质,能够切成不同厚度的连续切片,适合深埋于器官内部的组织或细胞的免疫细胞化学研究.介绍了应用Steedman's wax切片法观察植物细胞微管骨架的一般程序和方法以及经过作者检验且切实可行的一些技术改进.     

应用Steedman's wax 切片法观察植物细胞微管骨架

微管骨架在植物发育过程中起重要作用。由于植物细胞的特殊性,与动物细胞相比植物微管骨 架的研究遇到更多的困难。简略地介绍了曾被国内外学者应用的植物微管骨架的各种研究方法及其局限 性。Steedman's wax是一种多脂蜡。它熔点低(35~37℃),具有与石蜡相同的切片性质,能够切成不同厚 度的连续切片,适合深埋于器官内部的组织或细胞的免疫细胞化学研究。介绍了应用Steedman's wax 切 片法观察植物细胞微管骨架的一般程序和方法以及经过作者检验且切实可行的一些技术改进。     

Serial Sectioning of Insects with Hard Exoskeleton by Dissolution of the Exocuticle

The method reported here was designed to produce paraffin serial sections as thin as 5 Mm of insects or other arthropods with a hard cuticle. Heads and abdomens of Apis mellifera, Eristalomyia tenax and Tenebrio molitor were fixed with Schaffer's liquid, dehydrated with 80% ethanol, 90% ethanol, two changes of 100% isopropanol (2 hr each) and 12 hr in a 1:1 mixture of paraffin (58 C melting point) at 60 C. They were molded in paraffin after 12 hr of infiltration under a partial vacuum at 60 C. Large body openings of objects were sealed with paraffin to prevent infiltration of solvents.

Thereafter, the outer paraffin was removed manually and with xylene (15 min); the cuticle was rehydrated with 100% isopropanol and 100% ethanol (15 min each). The objects were then treated with Sputofluol (Merck; a mixture of NaOH and NaCIO) until they became white or their colorless endocuticle was stainable with aniline blue WS (C.I. 42755) after rinsing in a 50% acetic acid solution (v/v). They were then dehydrated with 100% ethanol and 100% isopropanol (15 min each) and subsequently re-embedded in paraffin. They were molded, sectioned, stained and mounted as usual.     

A Compact Co2 Chilling Device for Continuous Duty in Paraffin-Type Sectioning

To obtain uniform sections cut from some of the newer embedding materials of lower melting points, particularly the polyethylene glycols, it is necessary to cool the microtome knife. A cooling device constructed for this and other paraffin-type sectioning consists of a small brass wedge attached to the knife by means of an adjustable spring clip. The wedge is small enough to mount directly on the back of the knife and does not interfere with any movement of the microtome. Three small channels are drilled in the wedge to provide one entrance and two exits for gaseous CO₂. CO₂ from an upright tank is regulated by a modified Spencer Model 930 freezing attachment, which is connected to the brass wedge by copper capillary tubing.     

A Device for the Complete Elimination of Static Electricity in Paraffin Sectioning

In paraffin sectioning static electricity is often induced by friction between the knife and the block. The sections can be discharged by ionizing the air in the vicinity of the paraffin. This can be achieved by mounting a strip containing radioactive polonium near the microtome knife. This approach implies a certain radiological risk. An alternative solution is to apply a high electric field close to the knife. The latter method may offer hazards related to the high tension and production of ozone. In comparing the risks and benefits of the two methods we have concluded that the application of a high electric field offers less risks to people in the working area than the installation of a radioactive source. The costs of the two methods are of the same order.     

A New Method for Reducing Static Electricity During Paraffin Sectioning

Static electricity interferes with the production of good ribbons of thin paraffin sections. Sections tend to stick to the knife leading to compression, shredding and paraffin sections. Sections ribbon disintegration. the static electricity that builds up is caused by friction between the knife and the tissue block and by the rubbing together of the operator's clothing and sectioning table (Mattheij and Dignum 1975, Bryan and Hughes 1976).     

A New Method for Reducing Static Electricity During Paraffin Sectioning

Static electricity interferes with the production of good ribbons of thin paraffin sections. Sections tend to stick to the knife leading to compression, shredding and paraffin sections. Sections ribbon disintegration. the static electricity that builds up is caused by friction between the knife and the tissue block and by the rubbing together of the operator's clothing and sectioning table (Mattheij and Dignum 1975, Bryan and Hughes 1976).