Artefacts in the x-ray microanalysis of frozen-hydrated biological samples

1987 ◽  
Vol 45 ◽  
pp. 658-659
Author(s):  
Patrick Echlin
Keyword(s):  
Electron Scattering ◽  
Biological Samples ◽  
Fracture Face ◽  
Detailed Structure ◽  
X Ray ◽  
Morphological Appearance ◽  
The Poor ◽  
Freeze Dried

A number of papers have appeared recently which purport to have carried out x-ray microanalysis on fully frozen hydrated samples. It is important to establish reliable criteria to be certain that a sample is in a fully hydrated state. The morphological appearance of the sample is an obvious parameter because fully hydrated samples lack the detailed structure seen in their freeze dried counterparts. The electron scattering by ice within a frozen-hydrated section and from the surface of a frozen-hydrated fracture face obscures cellular detail. (Fig. 1G and 1H.) However, the morphological appearance alone can be quite deceptive for as Figures 1E and 1F show, parts of frozen-dried samples may also have the poor morphology normally associated with fully hydrated samples. It is only when one examines the x-ray spectra that an assurance can be given that the sample is fully hydrated.

Chemical modification of the bacterial wall to determine sites of metal deposition

1978 ◽  
Vol 36 (2) ◽  
pp. 350-351
Author(s):  
T. J. Beveridge
Keyword(s):  
Electron Scattering ◽  
Metal Salt ◽  
Metal Deposition ◽  
Suitable Model ◽  
X Ray Diffraction ◽  
Subtilis Cell ◽  
Thin Sections ◽  
X Ray ◽  
Section Data ◽  
Metal Impregnation

The Bacillus subtilis cell wall provides a protective sacculus about the vital constituents of the bacterium and consists of a collection of anionic hetero- and homopolymers which are mainly polysaccharidic. We recently demonstrated that unfixed walls were able to trap and retain substantial amounts of metal when suspended in aqueous metal salt solutions. These walls were briefly mixed with low concentration metal solutions (5mM for 10 min at 22°C), were well washed with deionized distilled water, and the quantity of metal uptake (atomic absorption and X-ray fluorescence), the type of staining response (electron scattering profile of thin-sections), and the crystallinity of the deposition product (X-ray diffraction of embedded specimens) determined.Since most biological material possesses little electron scattering ability electron microscopists have been forced to depend on heavy metal impregnation of the specimen before obtaining thin-section data. Our experience with these walls suggested that they may provide a suitable model system with which to study the sites of reaction for this metal deposition.

Revealing gross three-dimensional structure in the SEM

1987 ◽  
Vol 45 ◽  
pp. 656-657
Author(s):  
Sterling P. Newberry
Keyword(s):  
Freeze Drying ◽  
Three Dimensional ◽  
Dimensional Structure ◽  
Small Object ◽  
Final Solution ◽  
Dimensional Representation ◽  
X Ray ◽  
Three Dimensional Representation ◽  
Freeze Dried ◽  
Critical Point Drying

The beautiful three dimensional representation of small object surfaces by the SEM leads one to search for ways to open up the sample and look inside. Could this be the answer to a better microscopy for gross biological 3-D structure? We know from X-Ray microscope images that Freeze Drying and Critical Point Drying give promise of adequately preserving gross structure. Can we slice such preparations open for SEM inspection? In general these preparations crush more readily than they slice. Russell and Dagihlian got around the problem by “deembedding” a section before imaging. This some what defeats the advantages of direct dry preparation, thus we are reluctant to accept it as the final solution to our problem. Alternatively, consider fig 1 wherein a freeze dried onion root has a window cut in its surface by a micromanipulator during observation in the SEM.

Reduction of Potassium in Kidney Proximal Tubules to Aid in Electron Probe X-Ray Microanalysis of Calcium

1985 ◽  
Vol 43 ◽  
pp. 474-475
Author(s):  
A. LeFurgey ◽  
P. Ingram ◽  
L.J. Mandel
Keyword(s):  
Smooth Muscle ◽  
Proximal Tubule ◽  
Electron Probe ◽  
Clinical Applications ◽  
Proximal Tubules ◽  
Rabbit Kidney ◽  
Target Cells ◽  
X Ray ◽  
Freeze Dried

For quantitative determination of subcellular Ca distribution by electron probe x-ray microanalysis, decreasing (and/or eliminating) the K content of the cell maximizes the ability to accurately separate the overlapping K Kß and Ca Kα peaks in the x-ray spectra. For example, rubidium has been effectively substituted for potassium in smooth muscle cells, thus giving an improvement in calcium measurements. Ouabain, a cardiac glycoside widely used in experimental and clinical applications, inhibits Na-K ATPase at the cell membrane and thus alters the cytoplasmic ion (Na,K) content of target cells. In epithelial cells primarily involved in active transport, such as the proximal tubule of the rabbit kidney, ouabain rapidly (t1/2= 2 mins) causes a decrease2 in intracellular K, but does not change intracellular total or free Ca for up to 30 mins. In the present study we have taken advantage of this effect of ouabain to determine the mitochondrial and cytoplasmic Ca content in freeze-dried cryosections of kidney proximal tubule by electron probe x-ray microanalysis.

Electron Beam-Induced Mass Loss in Freeze-Dried Tissue and Protein Samples

1985 ◽  
Vol 43 ◽  
pp. 470-471
Author(s):  
M.E. Cantino ◽  
M.K. Goddard ◽  
L.E. Wilkinson ◽  
D.E. Johnson
Keyword(s):  
Electron Beam ◽  
Salivary Gland ◽  
Mass Loss ◽  
Counting Rate ◽  
Dose Dependence ◽  
X Ray ◽  
Large Samples ◽  
Freeze Dried ◽  
Muscle Homogenate ◽  
Large Muscle

Quantification in biological x-ray microanalysis depends on accurate evaluation of mass loss. Although several studies have addressed the problem of electron beam induced mass loss from organic samples (eg., 1,2). uncertainty persists as to the dose dependence, the extent of loss, the elemental constituents affected, and the variation in loss for different materials and tissues. in the work described here, we used x-ray counting rate changes to measure mass loss in albumin (used as a quantification standard), salivary gland, and muscle.In order to measure mass loss at low doses (10-4 coul/cm2 ) large samples were needed. While freeze-dried salivary gland sections of the required dimensions were available, muscle sections of this size were difficult to obtain. To simulate large muscle sections, frog or rat muscle homogenate was injected between formvar films which were then stretched over slot grids and freeze-dried. Albumin samples were prepared by a similar procedure. using a solution of bovine serum albumin in water. Samples were irradiated in the STEM mode of a JEOL 100C.

Preparation of cultured astrocytes for x-ray microanalysis

1989 ◽  
Vol 47 ◽  
pp. 988-989
Author(s):  
N.K.R. Smith ◽  
K.E. Hunter ◽  
P. Mobley ◽  
L.P. Felpel
Keyword(s):  
Cultured Cells ◽  
Elemental Content ◽  
Elemental Distribution ◽  
Sprague Dawley ◽  
X Ray ◽  
Cultured Astrocytes ◽  
Freeze Dried ◽  
Ultimate Objective

Electron probe energy dispersive x-ray microanalysis (XRMA) offers a powerful tool for the determination of intracellular elemental content of biological tissue. However, preparation of the tissue specimen , particularly excitable central nervous system (CNS) tissue , for XRMA is rather difficult, as dissection of a sample from the intact organism frequently results in artefacts in elemental distribution. To circumvent the problems inherent in the in vivo preparation, we turned to an in vitro preparation of astrocytes grown in tissue culture. However, preparations of in vitro samples offer a new and unique set of problems. Generally, cultured cells, growing in monolayer, must be harvested by either mechanical or enzymatic procedures, resulting in variable degrees of damage to the cells and compromised intracel1ular elemental distribution. The ultimate objective is to process and analyze unperturbed cells. With the objective of sparing others from some of the same efforts, we are reporting the considerable difficulties we have encountered in attempting to prepare astrocytes for XRMA.Tissue cultures of astrocytes from newborn C57 mice or Sprague Dawley rats were prepared and cultured by standard techniques, usually in T25 flasks, except as noted differently on Cytodex beads or on gelatin. After different preparative procedures, all samples were frozen on brass pins in liquid propane, stored in liquid nitrogen, cryosectioned (0.1 μm), freeze dried, and microanalyzed as previously reported.

Extended Core Edge Fine Structure in the E.M.

1980 ◽  
Vol 38 ◽  
pp. 120-121
Author(s):  
R.D. Leapman
Keyword(s):  
Fine Structure ◽  
Electron Scattering ◽  
Inelastic Electron ◽  
High Resolution Imaging ◽  
X Ray ◽  
Analytical Electron ◽  
Analytical Electron Microscope ◽  
Core Electrons ◽  
Resolution Imaging ◽  
X Ray Absorption

Extended X-ray Absorption Fine Structure (EXAFS) analysis makes use of synchrotron radiaion to measure modulations in the absorption coefficient above core edges and hence to obtain information about local atomic environments. EXAFS arises when ejected core electrons are backscattered by surrounding atoms and interfere with the outgoing waves. Recently, interest has also been shown in using inelastic electron scattering1-4. Some advantages of Extended X-ray-edge Energy Loss Fine Structure (EXELFS) are: a) small probes formed by the analytical electron microscope give spectra from μm to nm sized areas, compared with mm diameter areas for the X-ray technique, b) EXELFS can be combined with other techniques such as electron diffraction or high resolution imaging, and c) EXELFS is sensitive to low Z elements with K edges from ˜200 eV to ˜ 3000 eV (B to Cl).

scholarly journals Principles of Different X-ray Phase-Contrast Imaging: A Review

2021 ◽  
Vol 11 (7) ◽  
pp. 2971
Author(s):  
Siwei Tao ◽  
Congxiao He ◽  
Xiang Hao ◽  
Cuifang Kuang ◽  
Xu Liu
Keyword(s):  
Biological Samples ◽  
Phase Contrast ◽  
Recent Progress ◽  
Phase Contrast Imaging ◽  
Contrast Imaging ◽  
X Ray ◽  
X Ray Imaging ◽  
Internal Structures ◽  
X Ray Absorption ◽  
Made In

Numerous advances have been made in X-ray technology in recent years. X-ray imaging plays an important role in the nondestructive exploration of the internal structures of objects. However, the contrast of X-ray absorption images remains low, especially for materials with low atomic numbers, such as biological samples. X-ray phase-contrast images have an intrinsically higher contrast than absorption images. In this review, the principles, milestones, and recent progress of X-ray phase-contrast imaging methods are demonstrated. In addition, prospective applications are presented.

Sequential quantitative X-ray elemental imaging of frozen-hydrated and freeze-dried biological bulk samples in the SEM

2011 ◽  
Vol 245 (1) ◽  
pp. 17-25 ◽  
Author(s):  
ALAN T. MARSHALL ◽  
MELINDA J. GOODYEAR ◽  
SHEILA G. CREWTHER
Keyword(s):  
X Ray ◽  
Elemental Imaging ◽  
Freeze Dried

3D nanoscale imaging of biological samples with laboratory-based soft X-ray sources

2015 ◽  
Author(s):  
Aurélie Dehlinger ◽  
Anne Blechschmidt ◽  
Daniel Grötzsch ◽  
Robert Jung ◽  
Birgit Kanngießer ◽  
...  
Keyword(s):  
Biological Samples ◽  
X Ray ◽  
Nanoscale Imaging

scholarly journals Thermal, Structural, and Physical Properties of Freeze Dried Tropical Fruit Powder

2014 ◽  
Vol 2014 ◽  
pp. 1-10 ◽  
Author(s):  
K. A. Athmaselvi ◽  
C. Kumar ◽  
M. Balasubramanian ◽  
Ishita Roy
Keyword(s):  
Particle Size ◽  
Physical Properties ◽  
Average Particle Size ◽  
Average Particle ◽  
X Ray Diffraction ◽  
Tropical Fruit ◽  
X Ray ◽  
Freeze Dried ◽  
Fruit Powder ◽  
Lcr Meter

This study evaluates the physical properties of freeze dried tropical (guava, sapota, and papaya) fruit powders. Thermal stability and weight loss were evaluated using TGA-DSC and IR, which showed pectin as the main solid constituent. LCR meter measured electrical conductivity, dielectric constant, and dielectric loss factor. Functional groups assessed by FTIR showed presence of chlorides, and O–H and N–H bonds in guava, chloride and C–H bond in papaya, and chlorides, and C=O and C–H bonds in sapota. Particle size and type of starch were evaluated by X-ray diffraction and microstructure through scanning electronic microscopy. A semicrystalline profile and average particle size of the fruit powders were evidenced by X-ray diffraction and lamellar/spherical morphologies by SEM. Presence of A-type starch was observed in all three fruits. Dependence of electric and dielectric properties on frequency and temperature was observed.

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