An improved cryofixation technique for optimal orientation of Caenorhabditis elegans for cryosectioning and EPXMA

1995 ◽  
Vol 53 ◽  
pp. 1068-1069
Author(s):  
Edward Roberts ◽  
Nancy Wallace ◽  
Jonathan Freedman ◽  
Ann LeFurgey
Keyword(s):  
Caenorhabditis Elegans ◽  
Electron Probe ◽  
Intestinal Tract ◽  
Adult Stage ◽  
Elemental Distribution ◽  
Liquid Propane ◽  
X Ray ◽  
C Elegans ◽  
Optimal Orientation ◽  
Optimum Technique

INTRODUCTION: Plunge freezing into liquid cryogens has been frequently employed with success for cryofixation of biological specimens <lmm in size; however, positioning of cells or organisms with a precise orientation for subsequent cryosectioning is difficult. In these studies, a metal mirror cryofixation device, e.g. cryogun, has been tested to determine its applicability for cryofixation of small organisms such as nematodes or larval invertebrates with specific positioning. The objective of the experiments was to determine the optimum technique for rapid cryofixation and cryosectioning of the intestinal tract and lumen of the nematode Caenorhabditis elegans, prior to electron probe x-ray microanalysis (EPXMA) of subcellular elemental distribution.METHODS: Freezing and sectioning quality obtained using plunge freezing into -190°C liquid propane (Figure la) was compared with that obtained with -196°C metal mirror fixation using the cryogun (PS 1000, Delaware Diamond Knives, Wilmington, DE)(Figure lb). C. elegans were grown in culture to the adult stage, and individual nematodes were placed on either (1) a wooden specimen stub compatible with the plunge freezing device (Figure 2a) or (2) a cryogun specimen mount (Figure 2b).

Measurements of Morphology and Locomotion of Caenorhabditis Elegans With Digital Holographic Microscopy

2020 ◽  
Author(s):  
Yijie Wang ◽  
Jun Chen ◽  
Yuan Zhang ◽  
Kee-Hong Kim
Keyword(s):  
Caenorhabditis Elegans ◽  
Zebrafish Embryo ◽  
Adult Stage ◽  
Experimental Studies ◽  
Digital Holographic Microscopy ◽  
Development Stages ◽  
C Elegans ◽  
Applied Development ◽  
Time Period ◽  
Holographic Microscopy

Abstract Digital holographic microscopy (DHM) enables 3D volumetric measurements of small objects with high magnification. DHM has been applied to measure a variety of experimental studies, including turbulent boundary layer, spray droplets, individual cells, development of zebrafish embryo, etc. In this study, a DHM system is applied to measure the morphology and locomotion of two groups of Caenorhabditis Elegans (C. Elegans) with different development conditions (ATGL-1 group and n2 group) in an 8-day time period from their hatching to the adult stage, whose body lengths range from hundreds of micrometers to one millimeter. The length and volume are determined to describe the morphology of the C. Elegans at different development stages. The locomotion of the C. Elegans is divided into linear motion and curl motion. The kinetic energy derived from the two types of motion describes the extent of how active the C. Elegans is. The statistics of morphology and locomotion of the two groups of C. Elegans are compared at different development stages to illustrate the influence of the applied development conditions.

scholarly journals CROSSOVER SUPPRESSORS AND BALANCED RECESSIVE LETHALS IN CAENORHABDITIS ELEGANS

Genetics ◽  
1978 ◽  
Vol 88 (1) ◽  
pp. 49-65
Author(s):  
Robert K Herman
Keyword(s):  
Caenorhabditis Elegans ◽  
Linkage Group ◽  
The Other ◽  
Crossing Over ◽  
Linkage Groups ◽  
Recessive Lethal ◽  
X Ray ◽  
C Elegans ◽  
Complementation Groups ◽  
The Right

ABSTRACT Two dominant suppressors of crossing over have been identified following X-ray treatment of the small nematode C. elegans. They suppress crossing over in linkage group II (LGII) about 100-fold and 50-fold and are both tightly linked to LGII markers. One, called C1, segregates independently of all other linkage groups and is homozygous fertile. The other is a translocation involving LGII and X. The translocation also suppresses rrossing over along the right half of X and is homozygous lethal. CI has been used as a balancer of LGII recessive lethal and sterile mutations induced by EMS. The frequencies of occurrence of lethals and steriles were approximately equal. Fourteen mutations were assigned to complementation groups and mapped. They tended to map in the same region where LGII visibles are clustered.

Trauma causes heterogeneous compositional shifts in spinal cord

1992 ◽  
Vol 50 (2) ◽  
pp. 1594-1595
Author(s):  
Lyle G. Walsh ◽  
William B. Greene
Keyword(s):  
Spinal Cord ◽  
Electron Probe ◽  
Healthy People ◽  
Spinal Cord Trauma ◽  
Liquid Propane ◽  
X Ray ◽  
Cold Stage ◽  
Freeze Dried ◽  
Carbon Coated

Each year spinal cord trauma causes thousands of otherwise healthy people to be permanently disabled. In most cases, the axonal tracts are not mechanically severed. Instead, unknown mechanisms cause a progressive segmental necrosis. In this study, we use electron probe x-ray microanalysis, EPMA, to examine the composition of the dorsal axons and myelin 6 hours after mild and moderate trauma in order to identify the subcellular changes induced by trauma. Laminectomies were performed on anesthetized rats and sham, 6 g-cm or 20 g-cm trauma was delivered and the wound closed. Six hours after trauma the rats were again anesthetized, the dura was removed and the spinal cord was frozen in situ with a 100 psi jet of super-cooled liquid propane. Cryosections, 200 nm thick, were cut at -120°C and placed on carbon coated formvar covered folding copper grids. Samples were freeze dried and analyzed within an Hitachi H-7000 STEM on a Be tipped, GATAN, analytical cold stage with a 2-4 nA, 125 KeV beam.

Strategies For Combining Elemental Distribution Data Derived From Multiple Images and Samples

1998 ◽  
Vol 4 (S2) ◽  
pp. 134-135
Author(s):  
Marie E. Cantino ◽  
Joseph G. Eichen
Keyword(s):  
Electron Probe ◽  
Striated Muscle ◽  
Distribution Data ◽  
Elemental Distribution ◽  
Normalization Procedure ◽  
Multiple Images ◽  
X Ray ◽  
Freeze Dried ◽  
Hall Method ◽  
Elemental Peak

Data from digital electron probe x-ray microanalysis (EPXMA) images can be used to generate high resolution profiles of Ca binding within sarcomeres of vertebrate striated muscle. While ratios of elemental peak to bremstrahlung at each point (Hall method) have been used in many studies to allow comparison of data taken from different samples, this procedure has limitations for our application. A different normalization procedure is described here which provides a means of assessing variation among elemental and bremstrahlung profiles obtained from different images and samples.Freeze dried cryosections of chemically skinned frog or rabbit skeletal muscle were prepared as previously described. Digital EPXMA images were collected for 24 to 36 hours using a Zeiss EM910 STEM and an Oxford ExL2 microanalysis system. Calcium and bremstrahlung counts were summed within one pixel wide masks placed at successive positions along each half sarcomere, as previously described, using an automated routine in IPLAB on a Power Mac 7100.

The Investigation of Effects of Galvanizing Parameters on Microstructure of Al-Zn-Si Coating

2010 ◽  
Vol 154-155 ◽  
pp. 519-525
Author(s):  
Jia Shun Lv ◽  
Feng Li ◽  
Hong Gang Yang ◽  
Yong Lin Kang
Keyword(s):  
Surface Layer ◽  
Cooling Rate ◽  
Cross Section ◽  
Electron Probe ◽  
Solidification Process ◽  
Alloy Layer ◽  
Element Distribution ◽  
Elemental Distribution ◽  
X Ray Diffraction ◽  
X Ray

In this article, the effects of galvanizing process parameters, especially cooling rate, on the microstructures of coating was investigated. The microstructures of Al-Zn-Si coating were analyzed by the following methods: surface microstructure, cross-section microstructure, thickness and composition of alloy layer by SEM and its accessories, distribution of each element along depth by glow-discharge emission spectrometer, micro-area elemental distribution image analysis of each element distribution on surface and cross-section by electron probe, crystallization orientation by X-ray diffraction instrument. The results showed that coating was made up of aluminum-rich dendrite, interdendritic zinc-rich phase and alloy layer, that cooling rate affected depth of alloy layer in Al-Zn-Si coating directly, that zinc concentrated in the surface layer of coating and silicon in the alloy layer, while the coating has larger depth it also concentrated in the surface layer, that aluminum-rich dendrite has preferred orientation in the solidification process.

scholarly journals Characterization of aminopeptidase encoding gene anp-1 and its association with development in Caenorhabditis elegans

PeerJ ◽  
2019 ◽  
Vol 7 ◽  
pp. e7944 ◽  
Author(s):  
Shanchun Su ◽  
Baoliang Pan ◽  
Yanxin Hu ◽  
Ming Wang
Keyword(s):  
Caenorhabditis Elegans ◽  
Transcriptome Sequencing ◽  
Adult Stage ◽  
Small Body ◽  
Enrichment Analysis ◽  
Functional Expression ◽  
C Elegans ◽  
Sequencing Method ◽  
Differential Expression Genes ◽  
Development And Aging

Background Aminopeptidases play important roles in various biological processes in nematodes including growth, development and reproduction. Although the aminopeptidases have been shown to regulate reproduction in Caenorhabditis elegans (C. elegans), the role of aminopeptidases in development and aging has not been reported. This study focused on the function of aminopeptidase AlaNyl aminopeptidase 1 (ANP-1) on development in C. elegans. Methods In the present study, we reported the identification of ANP-1 in C. elegans along with sequence analysis and its functional expression and characterization. The phenotype changes were observed when anp-1 mutated. Then, differential expression genes (DEGs) between wild type strain (N2) and anp-1 deletion strain (RB804) were identified using transcriptome sequencing method. Finally, DEGs were verified by qRT-PCR assay. Results Our observations suggested that anp-1 mutation induced small body size in the L4/young adult stage of C. elegans, however, there was no difference between N2 and RB804 in adult stage. Moreover, deletion of anp-1 resulted in shortening lifespan and laying fewer eggs. DEGs (184 genes) were observed between N2 groups and RB804 groups by transcriptome sequencing. According to GO annotations and KEGG enrichment analysis, these DEGs play vital roles in development regulation in C. elegans. These data demonstrate ANP-1 participates in development and aging of C. elegans and will considerably contribute to the existing knowledge of aminopeptidase function in C. elegans.

Improvements in the electron probe forming capabilities and x-ray hole counts in the Philips TEM

1983 ◽  
Vol 41 ◽  
pp. 376-377
Author(s):  
Richard L. McConville
Keyword(s):  
Field Strength ◽  
Electron Probe ◽  
Spherical Aberration ◽  
Focal Length ◽  
Objective Lens ◽  
Parallel Beam ◽  
Tilt Axis ◽  
X Ray ◽  
Small Probe ◽  
Specimen Height

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.

Spatial resolution of X-ray microanalysis in thin foils

1978 ◽  
Vol 36 (1) ◽  
pp. 544-545
Author(s):  
R. Hutchings ◽  
I.P. Jones ◽  
M.H. Loretto ◽  
R.E. Smallman
Keyword(s):  
Spatial Resolution ◽  
Electron Probe ◽  
Beam Divergence ◽  
Inelastic Interaction ◽  
Specimen Thickness ◽  
High Current ◽  
Beam Spreading ◽  
X Ray ◽  
Thin Foils ◽  
Complex Calculation

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.

High spatial resolution x-ray microanalysis of interfaces

1995 ◽  
Vol 53 ◽  
pp. 284-285
Author(s):  
J. R. Michael
Keyword(s):  
Spatial Resolution ◽  
Electron Probe ◽  
Solid Angle ◽  
Collection Efficiency ◽  
Spatial Scales ◽  
Energy Dispersive Spectrometer ◽  
X Rays ◽  
X Ray ◽  
Probe Size ◽  
Brightness Field

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.

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