Mounting media, preparation

In all microscopic methods, sample preparation is key. Powder particles are normally dispersed in a mounting medium on a glass slide. Allen [7] has recommended that the particles not be mixed using glass rods or metal spatulas, as this may lead to fracturing a small camel-hair brush is preferable. A variety of mounting fluids with different viscosities and refractive indices are available a more viscous fluid may be preferred to minimize Brownian motion of the particles. Care must be taken, however, that the refractive indices of sample and fluid do not coincide, as this will make the particles invisible. Selection of the appropriate mounting medium will also depend on the solubility of the analyte [9]. After the sample is well dispersed in the fluid, a cover slip is placed on top... 

Finally, microscopic examination of samples often requires their preparation as cross sections or thin sections, or by mounting the sample on a glass slide by means of a mounting medium. For preparing thin and cross sections, samples are embedded in a polymer solution. After curing of the polymer, the thin or cross section is obtained by polishing the embedded sample with SiC abrasive disks. Aluminum suspensions or diamond paste are occasionally employed in a final polishing step. 

Small polished sections of the reaction products were prepared for reflecting-light microscopy by using Caulk Kadon or Technovit as a mounting medium. Identification of the phases either was undertaken in air or was aided by using oil immersion. In polished sections, depending on the optical properties of the phases examined, identification is often possible when as little as 0.01% of a phase is present. 

Slides are prepared by blending a fraction of a milligram of sample adhering to a paperclip or other clean disposable item into a drop of mounting medium on a slide and covering with a number 1% cover slip. Dipping the paperclip into the drop of mounting medium on the slide will make the particles adhere when the sample is touched. The particles should be spread in a thin even layer. 

A motility test is usefrd because B. anthracis is a nomnotile bacterium. Two motihty tests available are the wet mount and motihty medium variety. In a wet-mount preparation, organisms with Brownian movement or no movement will support the presence of B. anthracis. The presence of B. anthracis in a motdity medium preparation would be a single line of growth along the original inoculum stab (CDC, ASM, APHL, 2002). 

Mount the stained cells under a coverslip in a drop of the mounting medium and seal the preparation with melted paraffin or nail polish. The slides should be kept in the dark until measurement on LSC. 

Support Films. Since the specimens to be negative stained are generally in solution, some support medium is needed to mount the preparation on a grid for observation. While there are a number of choices in this regard, plastic and carbon are the two main compositional categories. 

Produce permanent preparations by mounting in an aqueous-based medium, such as Immu-mount, or in a xylene-based mounting medium, such as Fluoromount, following dehydration through an ethanol senes (i.e., 5 min each in 10, 20, 30, 40, 50, 60, 70, 80, and 90% ethanol in distilled water followed by three 5-min steps in absolute ethanol see Note 4). 

Section mounting medium 1% glycerin-albumin in distilled water, freshly prepared. 

Ono s method of cement kiln evaluation is based on observations of clinker or cement powder mounted in a liquid medium on a glass microscope slide. A polar ized-light microscope (the so called petrographic microscope) is an absolute necessity, and magniflca tions at approximately 400X are recommended. To determine the parameters of the kiln conditions in Ono s method as described in 1995, cement or clinker powder is sieved through a 100 pm screen (approxl mately U.S. Sieve No. 140), and a powder mount is prepared with a liquid of refractive index in the range of 1.705 to 1.715. The principal value of Ono s tech nique is that it can be employed by a competent, well trained microscopist on a small sample of clinker... 

Mowiol Mowiol, used as mounting medium, is prepared as described by Harlow and Lane (1988, p. 418) (see also article by Monika Herzog, Annette Draeger, Elisabeth Ehler, and J. Victor Small). Instead of DABCO we use 0.1% p-phenylenediamine as antifade compound. 

The selection of a suitable mounting compound is determined by the properties of the plastic, such as viscosity, shrinkage, adhesion to the sample, abrasion resistance, chemical resistance, and thermal resistance, as exhibited in the processes used to prepare the section and render the microstructure more visible. Another determining property is hardness, which serves as an indicator of possible edge rounding. For the most part, the requirements for a mounting medium are fulfilled by the plastics available on the market. 

To stain preparations with propidium iodide or TOTO-iodide, add these dyes to 90% glycerol, 2.5% n-propyl gallate mounting medium, to a final concentration of 1 dg/ml and 1/200, respectively (Gonzalez and Glover 1993 Carmena et al. 1998). 

Propidium iodide is a nucleic acid stain that is added to the mounting medium (for preparation, see p. 156 see also Figure 9.4A-C). Before staining with propidium iodide, the embryos must be treated with RNase to remove the RNA. 

Prepare mounting medium as described above. Add propidium iodide to a final concentration of 1 pg/ml. Store in the dark at -20°C. 

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