Specimen preparation method embedding

The optical microscope is used to study various fiber features, such as (1) size, (2) cross section (shape), (3) uniformity, (4) molecular orientation and (5) distribution of fillers. Specimen preparation methods include direct observation and sectioning. Fibers are embedded prior to sectioning by nucrotomy (Section 4.3) or polishing (Section 4.2) methods. Figure 5.1 contains optical... 

Cellulose nitrate and cellulose acetate (CA) were among the first asymmetric, reverse osmosis membranes to be produced [150]. Plummer et al [1511 described 13 specimen preparation methods for the observation of CA membrane structures. They pointed out the lack of contrast in epoxy embedded sections and that one of the best stains, osmium tetroxide, reacts with the... 

Cellulose nitrate and cellulose acetate (CA) were among the first asymmetric, reverse osmosis membranes to be produced [121]. Plummer et al. [122] described 13 specimen preparation methods for observation of CA membrane structures. They pointed out the lack of contrast in epoxy embedded sections and that one of the best stains, osmium tetroxide, reacts with the polymer. Freeze fractured membranes were found by these authors to be of questionable value. In our experience, if care is taken, SEM study of fractured membranes can provide an informative view of the structure even though some structures collapse, and their sizes cannot be accurately determined. A method found acceptable was ultrathin sectioning of gelatin embedded wet membranes (TEM). The structure of CA membranes was shown by replication [123] and SEM [124]. 

Conventional methods used for the preparation of biological material both for light microscopy and EM consist of fixation, dehydration, and embedding. These procedures result in increased permeability of the cell membrane with resultant loss or redistribution of mobile intracellular elements. Sections are then stained, resulting in the deposition of elements onto the specimen so that the resultant spectrum is not representative of the elemental composition in vivo (Figure 2). In consequence, such preparation methods are rarely used when the specimen is intended from the outset for microanalysis. 

Each view provides a different perspective on the membrane structure while, together, they give the complete structural model. Specimen preparation for OM and TEM cross sections was by nucrotomy of embedded membrane strips using a method developed to limit structural collapse (Section 4.3.4). An optical micrograph (Fig. 5.28A) shows the membrane cast on a woven support fabric. The active surface layer (top)... 

TTHIS CHAPTER IS FOR INDIVIDUALS who have not done electron microscopy (EM) before, but it also contains some new information about specimen preparation that can benefit experienced electron microscopists. It covers the materials and methods necessary to do routine thin sections of embedded specimens however, it does not cover specialized EM techniques such as freeze-fracture, rotary shadowing, negative staining, scanning EM, and serial sectioning. Likewise, this chapter focuses mainly on embryos because a description of the fixation methods for all the various types of fly tissues would require an entire book. This chapter is divided into three main sections ... 

The following procedure is more suitable for routine application than other methods as many as 200 specimens can be processed at a time with this procedure. Sections (2 pm thick) of formalin-fixed and paraffin-embedded tissues are mounted on silane-coated slides. They are deparaffmized with xylene and rehydrated in a series of descending concentrations of ethanol. The sections are immersed in 0.01 M sodium citrate buffer (pH 6.0) in plastic Coplin jars and heated in an autoclave at 120°C for 20min. After the sections have cooled down to room temperature for 20 min, they are incubated in the freshly prepared following silver staining solution for 25 min at room temperature. 

The usual method of sample preparation for tissue remains as formalin fixation and paraffin embedment (FFPE). This venerable approach may be satisfactory for the preservation of morphologic detail, but does adversely affect the antigenicity of many target molecules in the tissue, to degrees that are unknown. The enormous variation in protocols (including fixation times) employed for FFPE among different laboratories, or within the same laboratory from specimen to specimen, compounds the problem, and contributes to the current poor reproducibility. 

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