微波消解-石墨炉原子吸收光谱法测定大鼠组织中的微量硒

目的:建立微波消解石墨炉原子吸收光谱法(GFAAS)测定大鼠组织中的微量硒的检测方法.方法:采用微波消解法处理样品,用硝酸作为消解剂进行微波消解;采用氯化钯则作为基体改进剂来测定样品中的微量硒.结果:最佳灰化温度为900℃,原子化温度为2500℃;脑组织和肺组织的相对标准偏差(RSD)分别为2.83％和3.05％,样品加标回收率为97％～105％.结论:该方法用于检测大鼠组织中的硒含量,操作简便、快速、准确度好、结果满意,能够适用于生物体内多种组织器官中硒含量的测定与分析.     

一种快速、简易的扫描电镜植物样品干燥新方法

扫描电子显微镜是观察植物样品表面超微结构的有效方法,大部分新鲜植物样品需经过干燥处理才可以进行扫描电镜观察。该研究在传统叔丁醇冷冻干燥法的基础上,建立了叔丁醇一步冷冻干燥法,省略了固定、脱水、置换等步骤,简便易行,干燥后的样品形态饱满,最大程度保持了样品原貌。用叔丁醇一步冷冻干燥法干燥的样品可以与CO_2临界点干燥法和常规叔丁醇冷冻干燥法的效果相媲美。通过对不同的样品进行干燥处理,结果证明,该方法具有广泛适用性。     

禾本科植物叶片表皮气孔观察的样品制备方法改良

对现有的禾本科植物叶片气孔观察的样品制作方法中的不足作了一些改良。改良后的方法操作简便、耗时少、样品制备成功率高,且放大后的效果好,不会造成气孔形态的改变。改良方法适用于禾本科植物和其他叶肉紧实不易剥离的植物叶片。50%NaClO处理3min最适用于小麦旗叶表皮样品的制备。     

绒毡层及小孢子发育研究的扫描电镜样品制备

锇酸-二甲基亚砜一锇酸冷冻割断法制备的样品,在扫描电镜下,可观察绒毡层及小孢子发育过程,细胞形态保存良好,结构清晰,立体感强,为用扫描电镜研究植物细胞结构提供了一个有应用价值的方法。     

高脱乙酰度壳聚糖制备工艺研究

分别对传统的碱液加热法、超声波预处理法、微波辐射法三种制备壳聚糖的工艺进行了研究和比较。结果表明,以上三种方法均可以制备出脱乙酰度〉95％的壳聚糖,但利用微波辐射法制备壳聚糖所需生产设施少,工序简单,提高了效率,降低生产成本,环保节能,具有显著的技术先进性、经济性和实用性。     

绒毡层及小孢子发育研究的扫描电镜样品制备

锇酸-二甲基亚砜—锇酸冷冻割断法制备的样品,在扫描电镜下,可观察绒毡层及小孢子发育过程,细胞形态保存良好,结构清晰,立体感强,为用扫描电镜研究植物细胞结构提供了一个有应用价值的方法。     

植物游离细胞扫描电镜样品的制备法

介绍了植物游离细胞扫描电镜样品的制备方法步骤和注意事项.     

苯乙烯树脂割断法在生物样品制备中的应用

近年来样品割断法已成为扫描电镜样品制备的新技术之一.早在1968年Kachler已利用冰冻蚀刻复型技术,在透射电镜下观察了细胞内部结构.1970年Arenberg等对样品制备方法作了进一步改良,使扫描电镜也能观察细胞内部结构.1972年日本田中敬一设计了一种冷冻割断装置,观察组织块割断面上暴露出的各种管腔内面,以及割断面上细胞内的微细结构.目前冷冻割断技术已发展成树脂割断法、有机溶剂割断法、水溶性包埋剂割断法等多     

供光学显微镜观察的花粉样品制备的一种简单方法

传统的制备供光学显微镜观察的花粉样品的方法 (中国科学院植物研究所形态室孢粉组 ,1 960 )不但程序复杂 ,而且不同种类的花粉容易混杂。最近 ,我们在进行山茶属(Ｃａｍｅｌｌｉａ)花粉形态的系统研究中总结出一种制备供光镜观察的花粉材料的简单方法 ,现将其过程介绍如下 :( 1 )从标本或新鲜植株上取下花药 ,用冰醋酸浸软后 ,置洁净的凹玻片 (单凹玻片 )上 ,于解剖镜下将花药打开 ,滴上 95%酒精将花粉洗出。( 2 )滴上预先配制好的分解液 (醋酸酐 9份和浓硫酸 1份 ) ,于室温下或 50℃恒温箱里放置 5ｍｉｎ(具体温度和时间因花粉种类而异 ) …     

高水分、富含淀粉植物组织的扫描电镜制备技术优化

应用常规高真空扫描电子显微镜观察生物样品必须经过脱水和干燥处理,但无论采用临界点干燥还是冷冻干燥方法,都存在样品表面不同程度失真的问题。植物高水分、富含淀粉组织样品经处理后,容易出现淀粉流失、细胞壁变形等现象,从而造成扫描图像粗糙,无法获得真实的细胞内部结构。本文通过对CO_2临界点干燥、化学固定样品冷冻干燥和新鲜样品冷冻干燥3种扫描电镜样品制备技术中后期制样进行机械断裂和液氮脆断改进,优化出两种植物高水分、富含淀粉组织的扫描电镜样品制备方法:(1)样品首先进行FAA化学固定,经冷冻干燥后用液氮脆断,对断面喷金镀膜和扫描电镜观察。利用该方法所得细胞结构完整,细胞壁整齐,淀粉粒和蛋白轮廓明确,可用于分析淀粉粒和蛋白颗粒在细胞内的分布。(2)新鲜样品直接进行冷冻干燥,经液氮脆断后对断面喷金镀膜和扫描电镜观察。利用该方法所得细胞壁整齐,淀粉粒轮廓更清晰,并且无蛋白颗粒干扰,用于分析淀粉粒在细胞内的分布更加理想。     

Rapid cold fixation of tissue samples by microwave irradiation for use in electron microscopy

Summary A cold microwave irradiation procedure was developed to fix rapidly and stain various tissues and monolayers for electron microscopy. Because microwave stimulation always produces some heat, melting ice was used to maintain the temperature of the tissue samples, the fixative, and the staining solution at 0 to 4°C. The low temperature also reduced vapour formation, thus minimizing the risk of explosion. The microwave method shortened the total time of fixation and dehydration from the usual 3 h required by the conventional method to 65 min. After microwave fixation, the ultrastructural details of membranes and subcellular structures were excellent.     

微波快速免疫荧光组化染色方法的研究

本文应用微波辐射方法加速免疫荧光组化染色(间接和直接法),分别定位15种不同组织抗原,并应用连续切片同时用两种不同的孵育方法即微波辐射和常规孵育方法进行比较。结果证明,经微波辐射后免疫荧光组化染色时间大大缩短,背景染色明显好于常规法,阳性率和阳性强度与常规法基本一致。     

A COMPARISON OF METHODS FOR STOPPING INTERMEDIARY METABOLISM OF DEVELOPING RAT BRAIN1

Abstract— Freeze-blowing (Veech et al. 1973), focussed microwave irradiation (Stavinoha et al. 1973) and immersion in liquid nitrogen were compared as methods for stopping metabolism in order to assay in vivo levels of intermediary metabolites in developing rat brain. Freeze-blowing was superior at all ages (5. 10, 15 and 20 days post-natal). The differences between this method and immersion in liquid nitrogen were quite small in the youngest rats and increased with age. reflecting the increased time needed to freeze larger brains. Brains frozen by immersion in liquid nitrogen showed evidence of increased anaerobic metabolism, with increased fructose 1.6-diphosphate. dihydroxyacetone phosphate and lactate and decreased glucose 6-phosphate and creatine phosphate concentrations. When brain metabolism was stopped by microwave irradiation there were many differences from freeze-blown brain. Increases in fructose 1.6-diphosphate. dihydroxyacetone phosphate, ADP and AMP, and decreased in ATP and creatine phosphate were especially striking. The differences between microwave irradiation and freeze-blowing were not attributable simply to anoxia. Rather, the changes produced by this method seem to reflect the different thermal characteristics of the various enzymes which must be denatured to stop metabolism of the substrates measured. Unlike freezing in liquid nitrogen, the efficacy of microwave irradiation was not a simple function of head size, in that better results were achieved with 15- and 20-day-old than 5- or 10-day-old rats. Many glycolytic and Krebs cycle intermediates, as well as glutamate and aspartate, progressively increased over the course of development. The reasons for these increases are uncertain but are probably-related to the concomitant rises in rates of glycolysis and oxidative phosphorylation in brain.     

Microwave irradiation for a fast gas chromatography-mass spectrometric analysis of polysaccharide-based plasma volume expanders in human urine

In this contribution we tested the possibility to use microwave irradiation for the screening and confirmation pre-treatment steps of hydroxyethylstarch, with the aim to speed up gas chromatography-mass spectrometric procedures. Acid hydrolysis and derivatization processes were conducted in a temperature-controlled single beam microwave oven for organic synthesis. The kinetics of hydroxyethylstarch chemical hydrolysis and derivatization were investigated at different microwave power, incubation temperature and incubation time. The best hydrolysis conditions were found at a microwave power value of 1200 W (T 100°C) with an incubation time of 2 min; whereas the best derivatization conditions were found at a microwave power value of 1020 W (T 100°C) with an incubation time of 5 min. The effectiveness of this approach was evaluated by gas chromatography-mass spectrometry analyzing more than 20 different pools of blank urine samples spiked with hydroxyethylstarch at a concentration of 1 mg/mL. The results showed that the effect of microwave irradiation on the chemical hydrolysis process was very remarkable: the total sample preparation time can be shortened by 58 min compared to the reference method (2 min instead of 60 min). In addition to this, the time necessary for the derivatization process can also be drastically shortened with respect to the reference procedure (5 min instead of 30 min). The repeatability of the hydrolysis and derivatization recoveries, the limit of detection and the matrix interferences were comparable to the reference method accredited under the ISO 17025 guidelines and presently followed by the accredited sports anti-doping laboratory of Rome.     

THE ENZYMATIC MACERATION OF PLANT TISSUES: OBSERVATIONS USING A NEW METHOD OF MEASUREMENT

Mc Clendon , J. H., and G. F. Somers . (U. Delaware, Newark.) The enzymatic maceration of plant tissues: observations using a new method of measurement. Amer. Jour. Bot. 47(1) :1-7. Illus. 1960.—An apparatus is described for measuring the breaking strength of tissue slices. The apparatus was used in the measurement of maceration of potato tuber slices by fungal and tomato enzymes. During the enzymatic maceration of the slices, the strength fell in an approximately logarithmic manner to a stable value less than 5% of the initial strength. Calcium ion did not prevent enzymatic maceration, although it increased the strength of the tissue. Chelating agents used alone did not macerate but facilitated the enzymatic maceration. There was a pH optimum at 3.0—3.5 with a commercial “Pectinase” and enzymes from Botryosphaeria ribis but near 4.7 for a preparation from tomato fruit. The reciprocal of the time for a set strength reduction was proportional to the square root of the enzyme concentration. The relative strength remaining [(initial strength/final strength)–1] after an arbitrary reaction time was proportional to the enzyme concentration raised to the 0.8 power. The temperature coefficient was about 2.5, but other evidence indicated some limitation by diffusion. Non-enzymatic maceration increased rapidly below pH 3 and was especially prominent after subsequent neutralization.     

Microwave-assisted extraction of cyclotides from Viola ignobilis

Cyclotides are an interesting family of circular plant peptides. Their unique three-dimensional structure, comprising a head-to-tail circular backbone chain and three disulfide bonds, confers them stability against thermal, chemical, and enzymatic degradation. Their unique stability under extreme conditions creates an idea about the possibility of using harsh extraction methods such as microwave-assisted extraction (MAE) without affecting their structures. MAE has been introduced as a potent extraction method for extraction of natural compounds, but it is seldom used for peptide and protein extraction. In this work, microwave irradiation was applied to the extraction of cyclotides. The procedure was performed in various steps using a microwave instrument under different conditions. High-performance liquid chromatography (HPLC) and matrix-assisted laser desorption/ionization time-of-flight (MALDI–TOF) results show stability of cyclotide structures on microwave radiation. The influential parameters, including time, temperature, and the ratio of solvents that are affecting the MAE potency, were optimized. Optimal conditions were obtained at 20 min of irradiation time, 1200 W of system power in 60 °C, and methanol/water at the ratio of 90:10 (v/v) as solvent. The comparison of MAE results with maceration extraction shows that there are similarities between cyclotide sequences and extraction yields.     

TECHNIQUES FOR MEASURING VESSEL LENGTHS AND DIAMETERS IN STEMS OF WOODY PLANTS

Results were compared between the latex paint and compressed air methods for determining total vessel lengths, and between the sectioning and maceration methods for determining vessel diameters. The minimum, mean, median, and maximum vessel diameters were less with the sectioning method than with the maceration technique. Vessel diameter distributions were always nonnormal and had roughly similar patterns with the two techniques, but were statistically different from one another. In all six species where the paint and air methods for determining vessel length were compared, both methods showed a similar skewed vessel length distribution, with many short vessels and few long ones. Although there was no consistent pattern to the difference in results with these two methods, the vessel length frequency distributions were statistically different from one another. With the paint method, many vessels, especially many of the narrowest ones, were not paint-filled at the paint infusion port. The air method utilized the paint method, in part, and, in addition, is based upon the incorrect assumption that all vessels in the stem are the same diameter. Both techniques tended to exclude vessel lengths of the narrowest vessels. However, the narrow vessels, although numerous, contributed an insignificant amount to the total theoretical hydraulic conductance in stems.     

Does microwave irradiation have other than thermal effects on histological staining of the mammalian CNS? A light microscopical study of microwave stimulated staining under isothermal conditions in man and rat.

The question whether or not microwave irradiation exerts other than thermal effects on histological staining is still a matter of controversy. The present study was undertaken to reveal or reject such a so far hypothetical non-thermal irradiation effect. A device was developed, which enables exposure of histological sections or tissue pieces to microwave irradiation under isothermal conditions, i.e. with synchronous removal of the internal heat produced. Three classical neuroanatomical staining methods were tested on human and rat CNS. As control, identical procedures were performed without simultaneous microwave irradiation. The experiments were performed at three different temperature levels ranging from 5 to 50 degrees C. In none of the cases studied was a light microscopically appreciable difference observed between the microwave and non-microwave versions of a stain at the same temperature. The hypothesis of a separate non-thermal effect of microwave irradiation on histological staining is therefore rejected.     

Inactivation of Lactobacillus Bacteriophage PL-1 by Microwave Irradiation

The effect of microwave irradiation on the survival of bacteriophage PL-1, which is specific for Lactobacillus casei, was studied using a commercial 2,450 MHz microwave oven. The phages were inactivated by microwave irradiation according to almost first-order reaction kinetics. The rate of phage inactivation was not affected by the difference in the continuous or intermittent irradiation, nor by the concentrations of phages used, but was affected by the volume of phage suspensions, which prevented the loss of generated heat. Microwave irradiation of phage suspensions produced a number of ghost phages with empty heads, but fragmentation of the tail was hardly noticed. The breakage of phage genome DNA was primarily caused by the heat generated by microwave irradiation, whereas the phage DNA was not affected by the same temperature achieved by heat from outside. Thus we concluded that the phage-inactivating effect of microwave irradiation was mainly attributed to a thermal microwave effect, which was much stronger than a simple thermal exposure.     

Temperature oscillations in liquid media caused by continuous (nonmodulated) millimeter wavelength electromagnetic irradiation

Convection in liquids caused by 53–78 GHz millimeter wave irradiation with incident power density that ranged from 10 μW/cm² to 1 W/cm² was studied. Infrared thermography was used as an artifact-free method for recording surface-temperature dynamics during irradiation. It was found that continuous (nonmodulated) waves can produce a relaxation-type temperature oscillation in liquids with a relatively high stability of the period between temperature spikes. The temperature oscillation is due to the repetitive formation and dissipation of a torroidal type of convection vortex. When the vortex became stable during irradiation, we observed a temperature decrease following the initial temperature-rise phase, even though the irradiation was constantly maintained. This result constitutes a new process that can play a significant role in producing microwave bioeffects, including some so-called “nonthermal” effects and some effects that are inversely related to heating. Also, it can be considered as a newly discovered potential artifact in microwave bioeffects studies. © 1996 Wiley-Liss, Inc.