Effects of Various Fixing and Storage Fluids on Starch in Sapwood

Wood material that is to be used for observation of starch content may be stored many months in several plant material fixatives, formalin-acetic-alcohol, formalin-propionic-alcohol, tertiary butyl alcohol, ethyl alcohol, and 5 and 10% aqueous formalin without any apparent deleterious effect on the starch granules Storage of wood in aqueous formalin solutions of 15% and higher concentrations caused starch granules to break down and disperse forming a coagulated mass, in which it was impossible to see the individual grains.     

A Method for Embedding Thin Membranes in Historesin

A method is described for flat-embedding thin membranous tissues in Historesin. It allows easy orientation for sectioning large areas parallel to the surface. Selected fields can be monitored from the unfixed specimen, throughout preparation, to mounting on the microscope slide. For cross-sectioning, the flat-embedded tissue can be stacked and re-embedded to increase the amount of material examined per section.     

Thin Histological Sections Prepared from Large Thick Sections: a New Technique

A method is described for obtaining thin (1 μm) sections for light microscopy from large area thick (100 μm) sections of low viscosity nitrocellulose embedded specimens of human spinal osteoligamentous material.     

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.     

也门铁的组织培养技术研究

以4品种也门铁的茎段为外植体进行组织培养技术研究。结果表明,控制普通也门铁茎段褐化效果最理想的培养基为MS + 0.25 g·L-1 VC + 0.50 g·L-1 Na2S2O3;诱导也门铁不定芽萌动的最佳培养基为MS + 3.0 mg·L-1 6-BA + 0.2 mg·L-1 NAA + 0.1 mg·L-1 KT;诱导也门铁不定芽增殖的最佳培养基,因品种不同而异,普通也门铁和金心也门铁为MS + 2.0 mg·L-1 6-BA + 1.0 mg·L-1 NAA + 0.1 mg·L-1 GA3,扭纹铁和金心扭纹铁为MS + 1.0 mg·L-1 6-BA + 0.5 mg·L-1 NAA + 0.1 mg·L-1 GA3;也门铁壮苗的最佳培养基为MS + 0.05 mg·L-1 6-BA + 0.05 mg·L-1 NAA + l g·L-1AC;也门铁生根最优培养基为1/2MS + 0.5 mg·L-1 IBA。     

A tissue model for the study of cell proliferation in vitro

Summary A procedure for the cultivation of mesentery is described, in which the culture is fully representative of the tissue of origin. The intact mesenteric membrane—exposed to a minimum of trauma—was spread out over a hole in a filter paper strip in fluid medium and was cultivated free-hanging. Specimens from rats and guinea pigs were used. The organ culture model appears especially apt for cytochemical and proliferation studies. Proliferation variables based on Feulgen DNA analysis in individual, morphologically defined cells and on mitotic counting and radiochemical analysis were estimated. The tissue was fully viable in chemically defined growth medium and showed an almost unaltered light microscopical appearance after up to 52 hr in culture. Supported by the Swedish Cancer Society.     

A Plastic Embedding Technique for Analyzing Fluorescent Dextran-Amine Labelled Neuronal Profiles

A plastic embedding technique employing fluorescently labelled dextran-amines is described. After application of tracer to cut nerves and appropriate transport time, animals were fixed in paraformaldehyde. Subsequently their brains were dissected, heads and brains were dehydrated, embedded in methacrylate and sectioned serially on a rotary microtome. Plastic sections allow high resolution of single neuron profiles and complete serial reconstruction of un-distorted sections, including embryos with large amounts of yolk. In conjunction with whole mount analysis and double labelling, this technique can accurately reveal the spatial relationships of nerve components throughout development.     

A Method for Histological Examination of Undecalcified Teeth

A method is presented for histological examination of undecalcified ground sections of tooth roots affected with periodontal disease. The roots were placed in Karnovsky's fixative overnight, postfixed in 2% buffered osmic acid, and dehydrated in an ascending series of ethanol. The specimens were then infiltrated with propylene-oxide and Epon-Araldite resin, embedded in Epon-Araldite, and sections were prepared using a cutting and grinding system. The resulting ground sections were 8-12 μm thick. The sections were allowed to air dry at room temperature. When thoroughly dried, a coverglass was applied using resinous mounting medium DPX. The specimens were examined by phase-contrast microscopy. The method is useful for simultaneous examination of mineralized dental tissue and bacterial morphotypes covering the root surface of teeth involved with periodontal disease.     

A Method for Histological Examination of Undecalcified Teeth

A method is presented for histological examination of undecalcified ground sections of tooth roots affected with periodontal disease. The roots were placed in Karnovsky's fixative overnight, postfixed in 2% buffered osmic acid, and dehydrated in an ascending series of ethanol. The specimens were then infiltrated with propylene-oxide and Epon-Araldite resin, embedded in Epon-Araldite, and sections were prepared using a cutting and grinding system. The resulting ground sections were 8-12 μm thick. The sections were allowed to air dry at room temperature. When thoroughly dried, a coverglass was applied using resinous mounting medium DPX. The specimens were examined by phase-contrast microscopy. The method is useful for simultaneous examination of mineralized dental tissue and bacterial morphotypes covering the root surface of teeth involved with periodontal disease.     

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.     

A bright field/fluorescent stain for aluminum: its specificity,validation, and staining characteristics

A sensitive bright field/fluorescent histochemical staining method has been developed that reveals endogenous aluminum in subcellular structures. The method, achievable within 30 min, is based on phloxine B and phosphotungstic acid, with ethanol differentiation. Hematoxylin is used for nuclear and fast green FCF for cytoplasmic counterstaining. To test the method's specificity, we incubated living neuroblastoma cells overnight in culture media containing aluminum, calcium, iron, copper or zinc, or no added metal ions. After fixing the cells and applying the staining method, only cultures exposed to aluminum stained magenta. Applying the method to paraffin embedded tissue sections pretreated with one of two chelating agents that remove aluminum demonstrated less magenta staining in the chelated sections than in adjacent unchelated sections. Immersing sections overnight in solutions containing exogenous aluminum had no observable effect on staining for endogenous aluminum; therefore, it is unlikely that any exogenous aluminum present in histological reagents would alter the method's staining results.     

A bright field/fluorescent stain for aluminum: its specificity, validation, and staining characteristics

A sensitive bright field/fluorescent histochemical staining method has been developed that reveals endogenous aluminum in subcellular structures. The method, achievable within 30 min, is based on phloxine B and phosphotungstic acid, with ethanol differentiation. Hematoxylin is used for nuclear and fast green FCF for cytoplasmic counterstaining. To test the method's specificity, we incubated living neuroblastoma cells overnight in culture media containing aluminum, calcium, iron, copper or zinc, or no added metal ions. After fixing the cells and applying the staining method, only cultures exposed to aluminum stained magenta. Applying the method to paraffin embedded tissue sections pretreated with one of two chelating agents that remove aluminum demonstrated less magenta staining in the chelated sections than in adjacent unchelated sections. Immersing sections overnight in solutions containing exogenous aluminum had no observable effect on staining for endogenous aluminum; therefore, it is unlikely that any exogenous aluminum present in histological reagents would alter the method's staining results.     

组织工程研究的进展

组织工程从提出到现在不过10年的历史,但在世界范围内引起高度重视,许多国家已将它列为“经济的新增长点”,投入了相当的人力和财力进行研究,成果斐然.我国也有相关的研究课题开始运行.第二届组织工程学会于1998年在美国召开,会上发表了组织工程的最新研究成果.作为抛砖引玉,总结了有关组织工程研究的进展,并提出保证工程组织的功能化是值得深入研究的问题.     

Microperfusion Technique to Investigate Regulation of Microvessel Permeability in Rat Mesentery

Experiments to measure the permeability properties of individually perfused microvessels provide a bridge between investigation of molecular and cellular mechanisms regulating vascular permeability in cultured endothelial cell monolayers and the functional exchange properties of whole microvascular beds. A method to cannulate and perfuse venular microvessels of rat mesentery and measure the hydraulic conductivity of the microvessel wall is described. The main equipment needed includes an intravital microscope with a large modified stage that supports micromanipulators to position three different microtools: (1) a beveled glass micropipette to cannulate and perfuse the microvessel; (2) a glass micro-occluder to transiently block perfusion and enable measurement of transvascular water flow movement at a measured hydrostatic pressure, and (3) a blunt glass rod to stabilize the mesenteric tissue at the site of cannulation. The modified Landis micro-occlusion technique uses red cells suspended in the artificial perfusate as markers of transvascular fluid movement, and also enables repeated measurements of these flows as experimental conditions are changed and hydrostatic and colloid osmotic pressure difference across the microvessels are carefully controlled. Measurements of hydraulic conductivity first using a control perfusate, then after re-cannulation of the same microvessel with the test perfusates enable paired comparisons of the microvessel response under these well-controlled conditions. Attempts to extend the method to microvessels in the mesentery of mice with genetic modifications expected to modify vascular permeability were severely limited because of the absence of long straight and unbranched microvessels in the mouse mesentery, but the recent availability of the rats with similar genetic modifications using the CRISPR/Cas9 technology is expected to open new areas of investigation where the methods described herein can be applied.     

Effects of Inorganic Oxides,Sulfides, Chlorides,and Acid Chlorides on Metachromasia and Other Staining Properties

The ability of 17 inorganic compounds (POCl₃, PSC1₃, PC1₃, P₂O₅, P₂S₅, P₄S₃, P₄S₇, PC1₅, Sb₂O₅, As₂O₅, BiOC1₂, SeOC1₂, SO₂C1₂, Sb₂S₅, VOC1₂, SiC1₄ and CrO₂Cl₂) dissolved in pyridine or 2,2,4-trimethyl pentane, to enhance subsequent staining of tissue components with toluidine blue, phosphotungstic acid-hematoxylin (PTAH), leukofuchsin, and dihydroxydinaphthyl-disulfide (DDD) was studied. Eight of these compounds were also tested for ability to enhance staining with Alcian blue 8GN and Luxol fast blue MBS. Nine of the 17 compounds produced increased staining of certain tissue components with leukofuchsin, 13 with toluidine blue, 16 with PTAH, and 16 with DDD. The results suggest additional approaches to identification of tissue entities by induced metachromatic basophilia and leukofuchsin positivity as well as by the other stains studied, and also suggest a number of hitherto unstudied modes of reaction between the dyes used and reactive groups of tissue components. Many reactions of the compounds tested, with reactive groups known to be present in tissue components, are basecatalyzed, so that choice of solvent can influence the results obtained.     

A New Technique Facilitating Studies of Scant Cell Specimens

A simple method is described by which multiple cytological and cytochemical studies can be done on a clinical sample that contains relatively few cells. The cells are concentrated by centrifugation. The cell pellet is fixed, frozen and embedded in plastic. Thin (2-μm) sections are cut from the plastic. Thus, each cell may appear in several sections and many slides can be made from a single specimen. The advantages of this method over cytospins and Millipore filter preparations of cell suspensions are optimal utilization of all cells, excellent morphological and immunological preservation and ease and reproducibility of this technique.     

Fetal Tissue Banking for Transplantation: Characteristics of the Donor Population and Considerations for Donor and Tissue Screening

We initiated this study to evaluate the suitability for therapeutic use in transplantation of tissues obtained from human abortuses. We have developed protocols for the collection, handling and preservation of hepatic stem cells from electively aborted embryos and have developed methods for assessment of the cells so derived and processed. In this paper we present our findings regarding screening of potential donors, acquisition of fetal tissues, and assessment of the tissues for potentially infectious contaminants. We assess the suitability of the tissue donors according to current standards used for donors of commonly transplanted tissues (e.g., bone grafts, skin grafts and heart valves) and present data regarding the real availability of tissues from elective abortion procedures that would meet those standard tissue banking criteria.We specifically evaluated the donor's willingness to provide a blood sample for testing, conducted a detailed interview similar to those used for typical organ and tissue donors, and assessed the type and incidence of contamination in collected tissues. We find that although many women are willing to consent to use of the tissues for transplantation, attrition from the study for various reasons results in few fetal organs ultimately realistically available for transplantation. Typical reasons for attrition include: unwillingness to have a blood sample drawn or tested, positive serology results, social/medical high risk factors for acquisition of transmissible disease, no identifiable organs available, and unacceptable microbial contamination. Thus, although it might seem that due to the numbers of abortions performed annually, that there would be substantial numbers of suitable tissues available, only a small proportion are truly suitable for transplantation.     

A New Technique Facilitating Studies of Scant Cell Specimens

A simple method is described by which multiple cytological and cytochemical studies can be done on a clinical sample that contains relatively few cells. The cells are concentrated by centrifugation. The cell pellet is fixed, frozen and embedded in plastic. Thin (2-μm) sections are cut from the plastic. Thus, each cell may appear in several sections and many slides can be made from a single specimen. The advantages of this method over cytospins and Millipore filter preparations of cell suspensions are optimal utilization of all cells, excellent morphological and immunological preservation and ease and reproducibility of this technique.     

A simplified paraffin embedding method for small botanical samples

Abstract

We present a simplified paraffin embedding method suitable for unsuberized or unlignified small botanical samples (diameter < 0.3 cm). Only 2 h are required to yield plant tissues embedded in paraffin for anatomical observation and molecular analysis. Our method achieved morphological preservation of cell structures and conservation of nucleic acids that were equivalent to the traditional protocol. Fourier transform infrared spectrometry showed that the degree of degradation of the cytoplasmic components (e.g., protein) resulting from our simplified protocol was similar to that of the traditional protocol. The DNA samples embedded using the simplified method was extractable and could be used for PCR analysis. The DNA quality was equivalent to that embedded using the traditional method.