Specimen preparation for

Transmission electron microscopy is very widely used by biologists as well as materials scientists. The advantage of being able to resolve 0.2 nm outweighs the disadvantages of TEM. The disadvantages include the inabiUty of the common 100-kV electron beam to penetrate more than a few tenths of a micrometer (a 1000-kV beam, rarely used, penetrates specimens about 10 times thicker). Specimen preparation for the TEM is difficult because of the... 

Specimen Preparation for Transmission Electron Microscopy I. (J. C. Brav-man, R. M. Anderson, and M. L. McDonald, eds.) Volume 115 in MRS symposium proceedings series, 1988. 

S. Amelincks, D. van Dyck, J. van Landuyt, G. van Tendeloo (eds.) Electron Microscopy Principles and Fundamentals,VCH Verlagsgesellschaft mbH, Weinheim 1997. 2-178 R. M. Anderson, S. D. Walck (eds.) Specimen Preparation for Transmission Electron Microscopy of Materials IV, Materials Research Society, Pittsbrrrgh 1997. 

Brammar, 1. S. and Dewey, M. A. P., Specimen Preparation for Electron Metallography. Blackwell, Oxford (1966)... 

Moran DT, Rowley JC. Biological specimen preparation for correlative light and electron microscopy, in Correlative Microscopy in Biology, Instrumentation and Methods (Hayat MA, ed.), Academic Press, New York, 1987, pp. 2-22. 

With some exceptions (2-4), there have been relatively few recent reviews of microanalysis that have considered applications to plant science. In a previous review of this topic (5), I concentrated almost entirely on methods of specimen preparation for electron probe X-ray microanalysis. Here I highlight further developments in this area, and also broaden the scope of the review to include other microanalytical techniques. This chapter introduces the main types of hardware that are now available for microanalysis, reviews the main techniques used to prepare plant material prior to analysis, and provides protocols for the two major techniques. 

There is little doubt that cryotechniques, and particularly cryo-SEM, are now the dominant methods of specimen preparation for electron probe X-ray microanalysis when localization of soluble ions is required. In a previous review (5) these techniques were covered in considerable detail and this material is not reiterated here. Instead, protocols for the two major methods are provided and some recent developments and publications in this area are highlighted. 

J. Bravman, R. Anderson, and M. McDonald, Eds., Specimen preparation for transmission electron microscopy of materials (I), MRS Symp. Proc., Vol. 115, Pittsburgh, 1988. 

Glauert AM, Lewis PR (1998) Biological Specimen Preparation for Transmission Electron Microscopy. In Glauert AM (ed). Practical Methods in Electron Microscopy, vol 17. Portland Press, London... 

Csencsits, R. and Gronsky, R. (1988), in Specimen Preparation for Transmission Electron Microscopy of Materials MRS Symp. Proc., vol. 115 (eds ).C. Bravman, R.M. Anderson, and M.L McDonald), Materials Research Society, Pittsburg, PA, pp. 103-108. 

Metals, Pergamon Press, Oxford, 1959 P. A. Jacquet, Metallurgical Rev., 1,157 (1961) I. S. Brammer, M. A. P. Dewey, Specimen Preparation for Electron Metallography, American Elsevier, New York, 1966 J. W. Edington, Practical Electron Microscopy in Materials Science, Macmillan Philips Technical Library, Eindhoven, 1974. 

Causton, B. 1985. Does the embedding chemistry interact with tissue In Science of Biological Specimen Preparation for Microscopy and Microanalysis (M. Mueller, R. P. Becker, A. Boyde, and J. J. Wolosewick, eds.), pp. 209-214. SEM, Inc., AMF O Hare, IL. 

Fig. 2 Schematic of embedding and sectioning procedure used to limit relaxation of crack-tip microdeformation during specimen preparation for TEM (here for a CT specimen deformed under static loading)...

See, for example, the three-volume symposia proceedings entitled Specimen Preparation for Transmission Electron Microscopy of Materials, Vols. I—III, Materials Research Society, Warrendale, PA, 1987, 1990, and 1993. 

D. Bahnck and R. Hull, Experimental measurement of transmission electron microscope specimen temperature during ion milling, in R. Anderson, ed., Specimen Preparation for TEM, Vol. II, MRS Vol. 199, Materials Research Society, Warrendale, PA, 1990. 

Table I.—Specimen preparation for accelerated weathering, vertical-grained southern pine sapwood...

Figure 11.8. Illustration of sample preparation for cryo-TEM imaging. Parts (a)-(d) show specimen preparation for sample freezing (e) shows plunge freezing apparatus with temperature and humidity control and ( ) a typical image obtained by cryo-TEM. Copyright 2006 Nestec Etd.

The surface areas of dust samples as determined by optical and electron microscope have also been compared [167]. Pore size distributions of thin films of AI2O3, as measured by TEM, have also been compared with those determined by gas adsorption/desorption [168]. It has also been suggested that electron microscope gives a truer estimate of surface area than gas adsorption techniques [169]. Further information can be obtained in a recent review of specimen preparation for TEM [170]. 

Dykstra MJ, Reuss LE. Specimen Preparation for Electron Microscopy. 2003. Kluwer Academic/Plenum Publishers, New York. 

Specimen Preparation for SEM. In pharmaceutical technology, most of the samples that are examined by an SEM are powders. After the sampling procedure,... 

Watt, I.M. Specimen preparation for electron microscopy. In The Principle and Practice of Electron Microscopy, 2nd Ed. Cambridge University Press New York, 1997 136-187. 

The microstructure of a material can only be viewed in a light microscope after a specimen has been properly prepared. Metallurgists have developed extensive techniques and accumulated knowledge of metal specimen preparations for over a century. In principle, we can use these techniques to examine not only metallic materials but also ceramics and polymers in practice, certain modifications are needed and a certain degree of caution must be exercised. The main steps of specimen preparation for light microscopy include the following. 

Specimen preparation for cannabinoid testing frequently includes a hydrolysis step to free cannabinoids from their glucuronide conjugates. Most GC/MS confirmation procedures in urine measure total THCCOOH following either an enzymatic hydrolysis with //-glucuronidase, or more commonly, an alkaline hydrolysis with sodium hydroxide. Alkaline hydrolysis appears to efficiently hydrolyze the ester THCCOOH glucuronide linkage. 

Disadvantages of TEM depend on the researcher s point of view. Since the TEM operates under high vacuum conditions, live specimens cannot be imaged. In addition, specimen preparation for TEM is not trivial. In fact, in some cases, it is as much art as technology. In general, proper specimen preparation requires both auxiliary equipment and expertise. The more complex the research questions, the more complex and tedious the specimen preparation. This, however, can be approached - from a more positive stance. The vast capabilities of the TEM can be exploited by the selection of specific specimen preparation protocols that are suited to specific experimental designs. 

Specimen preparation for SPM differs in the inherent challenges depending on whether or not STM or AFM is used. One challenge common to both is the necessity of preparing a stable, fiat sample that is not pushed along the substrate by the probe. In addition, deformation of the sample by probe-tip interactions must be considered. 

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