The Use of Ethylenediamine in Softening Hard Plant Structures for Paraffin Sectioning

Ethylenediamine has been used as an agent for softening very hard woods prior to sectioning on a sliding microtome. The use of ethylenediamine is recommended for two additional uses: for preparing 1) soft woods in which wide, thin-walled tracheids or vessels tend to collapse during sliding microtome sectioning and 2) plant tissues with sclerenchyma mixed with soft-walled cells (bark, leaves, fruits, etc.) which frequently fail to section well. After softening in ethylenediamine, material is washed, infiltrated, and embedded in paraffin. Preliminary sections are made with a rotary microtome, just exposing the cut surface of the material; this exposed surface is soaked overnight in water. Sectioning is then continued. Sections produced in this fashion are considerably improved. The wood and pith of Podocarpus ustus, a parasitic conifer from New Caledonia, is used as an object to demonstrate improvements in sectioning by the ethylenediamine-paraffin method. Thinner sections with minimal tearing, cell collapse, and unevenness are produced. Sections can be handled easily and stained more effectively than unmounted sections. Variations in timing and in treatment are recommended to suit different materials. Ethylenediamine, used with reasonable caution, is much less hazardous than hydrofluoric acid and is more effective in softening plant material. The ethylenediamine method may be used routinely on any material difficult to section because of hardness.     

Isopropyl Alcohol in the Paraffin Infiltration Technic

A method is described for using isopropyl alcohol for dehydration of animal tissues preceding melted paraffin infiltration. Advantages of the technic are: simplicity, low cost, low toxicity, and diminished distortion and hardening of the tissues.     

植物材料快速石蜡制片方法

真空干燥箱已越来越广泛地应用于现代生物学研究领域。该文利用真空干燥箱温度和负压的可控制性能,将固定、脱水、透明和石蜡渗透等过程在真空干燥箱中进行,建立起一套可行的植物组织快速石蜡制片方法。结果显示,真空干燥箱的应用加速了多种试剂的渗透速率,提高了切片质量,达到了优化实验步骤、节省实验时间和减少室内有毒化学气体污染的目的。     

A Rigid Illuminator Giving Transmitted Light to Facilitate Microhardness Testing of Calcified Tissues

The structural form of calcified tissues necessitates their examination with transmitted light to identify various morphological features which are not evident with incident illumination alone. To achieve stability for indentation, the specimens must be rigidly mounted. A Leitz Model 514095 base illuminator was fitted with a bottom plate of 3 mm brass for attaching it to the graduated stage and a perforated, 3 mm thick brass top plate, surmounted by a 6 mm thick piece of plate glass. The 25 mm hole in the top plate coincided with a ground glass disc to transmit diffused light through the plate glass to the specimen. Thus, the specimen could be examined on a rigid microscope stage, morphological features identified, and subsequent interpretation of indents made with the usual incident illumination.     

Histochemical Demonstration of Acid Phosphatase in Hard Tissues

The action of the following decalcifying solutions for the demonstration of acid phosphatase has been studied: buffer solution acetic-acetate 0.05 M, pH 5; 2, 5, 10, 20, and 50% formic acid and 20% sodium citrate in equal parts (pH 2.6, 3, 3.8, 4.2, and 5); 0.5 M citric acid-NaOH, pH 4.2; Versene solution, 5%, pH 7. A comparative study of fixatives has been made also (neutral formalin, 10-20%; formalin-chloral hydrate (Fishman), acetone and 80% alcohol). The best results were obtained with fixation at 4°C in 10-20% neutral formalin or formalin-chloral hydrate, for a period of 24 hr, and decalcification with 20% sodium citrate, 5% formic acid, in equal parts, pH 4.2, which can act on both specimens or sections for a period up to 2 wk with very little loss of enzyme. It is not necessary to reactivate the enzyme after decalcification; frozen sections should be used and should be washed in distilled water before proceeding with the demonstration of the enzyme (Gomori's method or azo dye coupling). Other fixatives (acetone and alcohol) and paraffin embedding produce a greater loss in enzymes and very irregular results.     

Softening Hair with Dithiothreitol (Cleland's Reagent) for Histological Sectioning in Paraffin

Undecalcified embedment of large bone specimens is often challenging. A method is presented here that is suitable for methacrylate embedment of sections of canine vertebrae while retaining the ability to localize tartrate-resistant acid phosphatase and alkaline phosphatase activity. Specimens also retained tetracycline labelling, and sectioned preparations were readily stained with routine bone procedures. A modification of the Bodian silver stain, used for examining the nerves and spinal cord in these specimens, provided a useful stain for canaliculi and cement lines in trabecular and cortical bone. This stain is advantageous when both bone and nerve tissue are of interest, as in spinal fusion studies.     

Elimination OF Distortion and Poor Staining in Paraffin Sections OF Plant Tissues

Three fixing solutions causing least distortion and bright staining of plant tissues are named. Glycerin dehydration causes less distortion than a series of alcohol concentrations; 95% alcohol removes some of the glycerin, sets the protoplasm and improves the staining. Absolute alcohol causes distortion and should be avoided. Pure chloroform, as a paraffin solvent, is followed by brighter staining but more distortion than are the butyl alcohols. A schedule resulting in minimum distortion is given. The results are shown in photomicrographs. Brightest staining follows the use of C. P. iron alum and hematoxylin. The use of a paper cup for very gradual change from one liquid to another and as a labor saver is described.     

Gold Toning Improves the Visualization of Nucleolar Organizer Regions in Paraffin Embedded Tissues

A modification of the silver colloid technique for staining nucleolar organizer regions in paraffin embedded tissues is described. This modification involves the application of a gold toning step with subsequent gold reduction, if necessary, following incubation of sections in the standard silver colloid solution. Silver stained nucleolar organizer regions (AgNORs) in toned sections are more sharply delineated when compared to untoned controls. in high grade tumors the addition of the toning step results in significantly higher AgNOR counts due to the ability to discriminate more easily individual AgNORs in argyrophilic aggregates within the nucleus. It is recommended, because of enhanced visualization, that this modification of the silver colloid technique be used in studies involving quantification of AgNORs in tissue sections.     

Gold Toning Improves the Visualization of Nucleolar Organizer Regions in Paraffin Embedded Tissues

A modification of the silver colloid technique for staining nucleolar organizer regions in paraffin embedded tissues is described. This modification involves the application of a gold toning step with subsequent gold reduction, if necessary, following incubation of sections in the standard silver colloid solution. Silver stained nucleolar organizer regions (AgNORs) in toned sections are more sharply delineated when compared to untoned controls. in high grade tumors the addition of the toning step results in significantly higher AgNOR counts due to the ability to discriminate more easily individual AgNORs in argyrophilic aggregates within the nucleus. It is recommended, because of enhanced visualization, that this modification of the silver colloid technique be used in studies involving quantification of AgNORs in tissue sections.     

Structure and Stress-Strain Relationship of Soft Tissues

The mechanical properties of a soft tissue are related to itsstructure. Weshall illustrate this by the properties of thearteries and the lung. Viscoelasticity, strain rate effects,pseudo-elasticity, and constitutive equations ar discussed.The mecahnical properties of an organ is, however, not onlybased onthe tissues of the organ, but also on its geometry andrelationship to the neighboring organs. A typical example isthe blood vessel. The capillary blood vessels of the mesenteryare "rigid"; those in the bat's wing are "distensible"; whereasthe capillaries of the lung are "sheet" like: rigid in one plane,and compliant in another. The stress-strain relationship ofthe systemic arteries is highly nonlinear, stiffening exponentiallywith increasing strains; yet that of the pulmonary arteriesin the lung is linear. The systemic veins are easily collapsible;yet the pulmonary veins in the lung are not: they remain patentwhen the blood pressure falls below the alveolargas pressure.The explanation of these differences lies in the varied interactionsbetween the blood vessels and the surrounding tissues in differentorgans. The implications of these differences on blood circulationare pointed out. Therole of ultrastructure is discussed.     

Roentgen Examination of Soft Tissues of the Pelvis

With meticulous preparation of the patient and with careful technique, the soft tissues of the pelvis are identifiable in most cases. Search should be made for the traces of abnormal pelvic structures on plain-film studies. Once the normal is recognized, any variations are easily identified. The fundamental differences between various radiologic densities—air, fat, fluid, muscle, calcium, bone and metal—should be observed. Special procedures can be used to enhance the contrasts after adequate evaluation of the simplest and, on many occasions, the invaluable, plain-film study of the soft tissues of the pelvis.     

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.     

Staining of Animal Tissues with the Dye Base of Methylene Green in Benzene to Facilitate Identification and Selection of Material

Methylene green as the free dye base dissolved in benzene can be used to stain animal tissues sufficiently during clearing for paraffin embedding to distinguish morphological features. It can therefore be used to allow trimming off unwanted tissues, to orient specimens when casting in wax and to select sections from a wax ribbon. The stain is removed very quickly on contact with water, and has no effect on subsequent staining techniques. The dye base is prepared by adding 2.5 ml of 10% NaOH to 100 ml of 1% aqueous methylene green, mixing well and allowing to stand at 20-30 C for 24 hr. The precipitate is separated by filtration, washed on the paper and dried at 50-55 C. The dry precipitate is dissolved in 200 ml of benzene to make the stock solution, which is diluted with 1 or 2 parts of benzene for tissue blocks, but used undiluted on sections.     

Er-YAG激光在口腔软组织中的应用

随着科学技术的进步,激光技术正以惊人的速度向前发展。自激光引入口腔治疗中以来,被广泛的应用于口腔硬组织中。由于激光杀菌、消毒等特性研究的广泛报道,近来激光应用于口腔软组织也得到了越来越多的关注。其中Er-YAG激光在口腔领域的实用性和安全性已得到多方面的认证。该文就激光在口腔医学特别是软组织中的应用作一综述。     

A Novel Nonlinear Parameter Estimation Method of Soft Tissues

The elastic parameters of soft tissues are important for medical diagnosis and virtual surgery simulation. In this study, we propose a novel nonlinear parameter estimation method for soft tissues. Firstly, an in-house data acquisition platform was used to obtain external forces and their corresponding deformation values. To provide highly precise data for estimating nonlinear parameters, the measured forces were corrected using the constructed weighted combination forecasting model based on a support vector machine(WCFM_SVM). Secondly, a tetrahedral finite element parameter estimation model was established to describe the physical characteristics of soft tissues,using the substitution parameters of Young's modulus and Poisson's ratio to avoid solving complicated nonlinear problems. To improve the robustness of our model and avoid poor local minima,the initial parameters solved by a linear finite element model were introduced into the parameter estimation model. Finally, a self-adapting Levenberg–Marquardt(LM) algorithm was presented,which is capable of adaptively adjusting iterative parameters to solve the established parameter estimation model. The maximum absolute error of our WCFM_SVM model was less than 0.03 Newton, resulting in more accurate forces in comparison with other correction models tested.The maximum absolute error between the calculated and measured nodal displacements was less than 1.5 mm, demonstrating that our nonlinear parameters are precise.