Cytochemical and electron probe X-ray microanalysis studies on the distribution change of intracellular calcium in columella cells of soybean roots under simulated microgravity

2011 ◽  
Vol 61 (1) ◽  
pp. 57-69 ◽  
Author(s):  
M. Hayatsu ◽  
M. Ono ◽  
C. Hamamoto ◽  
S. Suzuki
Author(s):  
Minoru Onozuka ◽  
Eiichi Sugaya ◽  
Aiko Sugaya ◽  
Masayoshi Usami

The role of divalent cations, particularly of calcium, has gained increasing interest as a charge carrier and in processes such as regulation of enzymatic activities, in secretions of humoral transmitters and initiation of muscle contraction. The role of calcium has become very important in manifesting the bursting activity of neurons by various electrophysiological techniques, especially by voltage clamping. The distribution of calcium compared with the ultrastructure of nerve cells, however, has not been widely investigated. Analysis by the electron probe X-ray microanalyser(EPXMA) makes this type of research easier. To determine the relationship between calcium localization within the neuron and cellular function, we tried to make an intracellular calcium distribution map of the normal state and that during bursting activity by the computer controlled EPXMA. We also detected the chemical shift between the different states of cellular function caused by the intracellular calcium binding state.


Author(s):  
Richard L. McConville

A second generation twin lens has been developed. This symmetrical lens with a wider bore, yet superior values of chromatic and spherical aberration for a given focal length, retains both eucentric ± 60° tilt movement and 20°x ray detector take-off angle at 90° to the tilt axis. Adjust able tilt axis height, as well as specimen height, now ensures almost invariant objective lens strengths for both TEM (parallel beam conditions) and STEM or nano probe (focused small probe) modes.These modes are selected through use of an auxiliary lens situ ated above the objective. When this lens is on the specimen is illuminated with a parallel beam of electrons, and when it is off the specimen is illuminated with a focused probe of dimensions governed by the excitation of the condenser 1 lens. Thus TEM/STEM operation is controlled by a lens which is independent of the objective lens field strength.


Author(s):  
R. Hutchings ◽  
I.P. Jones ◽  
M.H. Loretto ◽  
R.E. Smallman

There is increasing interest in X-ray microanalysis of thin specimens and the present paper attempts to define some of the factors which govern the spatial resolution of this type of microanalysis. One of these factors is the spreading of the electron probe as it is transmitted through the specimen. There will always be some beam-spreading with small electron probes, because of the inevitable beam divergence associated with small, high current probes; a lower limit to the spatial resolution is thus 2αst where 2αs is the beam divergence and t the specimen thickness.In addition there will of course be beam spreading caused by elastic and inelastic interaction between the electron beam and the specimen. The angle through which electrons are scattered by the various scattering processes can vary from zero to 180° and it is clearly a very complex calculation to determine the effective size of the beam as it propagates through the specimen.


Author(s):  
J. R. Michael

X-ray microanalysis in the analytical electron microscope (AEM) refers to a technique by which chemical composition can be determined on spatial scales of less than 10 nm. There are many factors that influence the quality of x-ray microanalysis. The minimum probe size with sufficient current for microanalysis that can be generated determines the ultimate spatial resolution of each individual microanalysis. However, it is also necessary to collect efficiently the x-rays generated. Modern high brightness field emission gun equipped AEMs can now generate probes that are less than 1 nm in diameter with high probe currents. Improving the x-ray collection solid angle of the solid state energy dispersive spectrometer (EDS) results in more efficient collection of x-ray generated by the interaction of the electron probe with the specimen, thus reducing the minimum detectability limit. The combination of decreased interaction volume due to smaller electron probe size and the increased collection efficiency due to larger solid angle of x-ray collection should enhance our ability to study interfacial segregation.


Author(s):  
P. Ingram

It is well established that unique physiological information can be obtained by rapidly freezing cells in various functional states and analyzing the cell element content and distribution by electron probe x-ray microanalysis. (The other techniques of microanalysis that are amenable to imaging, such as electron energy loss spectroscopy, secondary ion mass spectroscopy, particle induced x-ray emission etc., are not addressed in this tutorial.) However, the usual processes of data acquisition are labor intensive and lengthy, requiring that x-ray counts be collected from individually selected regions of each cell in question and that data analysis be performed subsequent to data collection. A judicious combination of quantitative elemental maps and static raster probes adds not only an additional overall perception of what is occurring during a particular biological manipulation or event, but substantially increases data productivity. Recent advances in microcomputer instrumentation and software have made readily feasible the acquisition and processing of digital quantitative x-ray maps of one to several cells.


Author(s):  
A. LeFurgey ◽  
P. Ingram ◽  
L.J. Mandel

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.


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