Structural analysis of crystalline Ca++ transport ATPase vesicles

1983 ◽  
Vol 41 ◽  
pp. 636-637
Author(s):  
E. Loren Buhle ◽  
Barry E. Knox ◽  
Ueli Aebi
Keyword(s):  
Electron Microscopy ◽  
Sarcoplasmic Reticulum ◽  
Three Dimensional ◽  
Integral Membrane Proteins ◽  
Molecular Organization ◽  
Dimensional Structure ◽  
Crystal Formation ◽  
Transport Atpase ◽  
Ordered Arrays ◽  
Tubular Arrays

In our efforts to study the three-dimensional structure and molecular organization of ion-motive ATPases by electron microscopy and image processing we have systematically investigated conditions which induce the formation of ordered arrays of these integral membrane proteins. Among them we have explored the effect(s) of vanadate and other multi-valent ions on ATPase crystal formation by starting from published procedures [e.g. 1,2]. Here we present some preliminary results concerned with the packing and molecular organization of vanadate-induced crystalline arrays of a Ca++ transport ATPase in tubular rabbit sarcoplasmic reticulum vesicles.The Ca++ activated ATPase from rabbit sarcoplasmic reticulum was purified as published. Induction of ordered tubular arrays of the protein with vanadate was basically achieved as described, except that the protein was slowly dialysed against a vanadate-containing buffer rather than directly adding the vanadate to the protein. Optimal yields of crystalline arrays were obtained after two days of dialysis at 4°C.

Electron Microscopy of Cytoplasmic Contractile Proteins

1978 ◽  
Vol 36 (3) ◽  
pp. 606-614 ◽  
Author(s):  
T.D. Pollard ◽  
P. Maupin
Keyword(s):  
Electron Microscopy ◽  
Actin Filaments ◽  
Three Dimensional ◽  
Uranyl Acetate ◽  
Contractile Proteins ◽  
Dimensional Structure ◽  
X Ray Diffraction ◽  
Cytoplasmic Matrix ◽  
Computer Image Processing ◽  
Nonmuscle Cells

In this paper we review some of the contributions that electron microscopy has made to the analysis of actin and myosin from nonmuscle cells. We place particular emphasis upon the limitations of the ultrastructural techniques used to study these cytoplasmic contractile proteins, because it is not widely recognized how difficult it is to preserve these elements of the cytoplasmic matrix for electron microscopy. The structure of actin filaments is well preserved for electron microscope observation by negative staining with uranyl acetate (Figure 1). In fact, to a resolution of about 3nm the three-dimensional structure of actin filaments determined by computer image processing of electron micrographs of negatively stained specimens (Moore et al., 1970) is indistinguishable from the structure revealed by X-ray diffraction of living muscle.

High-voltage electron microscopy of maxillary gland of spirochete-infected brine shrimp: lesion of efferent tubule

1988 ◽  
Vol 46 ◽  
pp. 362-363
Author(s):  
G. E. Tyson ◽  
M. J. Song
Keyword(s):  
Electron Microscopy ◽  
High Voltage ◽  
Brine Shrimp ◽  
Natural Populations ◽  
Three Dimensional ◽  
Dimensional Structure ◽  
High Voltage Electron Microscopy ◽  
Voltage Electron ◽  
High Voltage Electron ◽  
Infected Adult

Natural populations of the brine shrimp, Artemia, may possess spirochete- infected animals in low numbers. The ultrastructure of Artemia's spirochete has been described by conventional transmission electron microscopy. In infected shrimp, spirochetal cells were abundant in the blood and also occurred intra- and extracellularly in the three organs examined, i.e. the maxillary gland (segmental excretory organ), the integument, and certain muscles The efferent-tubule region of the maxillary gland possessed a distinctive lesion comprised of a group of spirochetes, together with numerous small vesicles, situated in a cave-like indentation of the base of the tubule epithelium. in some instances the basal lamina at a lesion site was clearly discontinuous. High-voltage electron microscopy has now been used to study lesions of the efferent tubule, with the aim of understanding better their three-dimensional structure.Tissue from one maxillary gland of an infected, adult, female brine shrimp was used for HVEM study.

Confocal laser microscopy of impregnated central nervous system tissue

1988 ◽  
Vol 46 ◽  
pp. 94-95
Author(s):  
J.N. Turner ◽  
M. Siemens ◽  
D. Szarowski ◽  
D.N. Collins
Keyword(s):  
Electron Microscopy ◽  
Central Nervous System ◽  
Nervous System ◽  
Three Dimensional ◽  
Dimensional Structure ◽  
Confocal Laser ◽  
High Contrast ◽  
Central Nervous System Tissue ◽  
Tissue Samples ◽  
Reflected Light

A classic preparation of central nervous system tissue (CNS) is the Golgi procedure popularized by Cajal. The method is partially specific as only a few cells are impregnated with silver chromate usualy after osmium post fixation. Samples are observable by light (LM) or electron microscopy (EM). However, the impregnation is often so dense that structures are masked in EM, and the osmium background may be undesirable in LM. Gold toning is used for a subtle but high contrast EM preparation, and osmium can be omitted for LM. We are investigating these preparations as part of a study to develop correlative LM and EM (particularly HVEM) methodologies in neurobiology. Confocal light microscopy is particularly useful as the impregnated cells have extensive three-dimensional structure in tissue samples from one to several hundred micrometers thick. Boyde has observed similar preparations in the tandem scanning reflected light microscope (TSRLM).

scholarly journals Capturing Enveloped Viruses on Affinity Grids for Downstream Cryo-Electron Microscopy Applications

2013 ◽  
Vol 20 (1) ◽  
pp. 164-174 ◽  
Author(s):  
Gabriella Kiss ◽  
Xuemin Chen ◽  
Melinda A. Brindley ◽  
Patricia Campbell ◽  
Claudio L. Afonso ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Measles Virus ◽  
Influenza A ◽  
Electron Tomography ◽  
Three Dimensional ◽  
Nitrilotriacetic Acid ◽  
Dimensional Structure ◽  
Enveloped Viruses ◽  
Cryo Electron Microscopy ◽  
Human Immunodeficiency Virus Virus

AbstractElectron microscopy (EM), cryo-electron microscopy (cryo-EM), and cryo-electron tomography (cryo-ET) are essential techniques used for characterizing basic virus morphology and determining the three-dimensional structure of viruses. Enveloped viruses, which contain an outer lipoprotein coat, constitute the largest group of pathogenic viruses to humans. The purification of enveloped viruses from cell culture presents certain challenges. Specifically, the inclusion of host-membrane-derived vesicles, the complete destruction of the viruses, and the disruption of the internal architecture of individual virus particles. Here, we present a strategy for capturing enveloped viruses on affinity grids (AG) for use in both conventional EM and cryo-EM/ET applications. We examined the utility of AG for the selective capture of human immunodeficiency virus virus-like particles, influenza A, and measles virus. We applied nickel-nitrilotriacetic acid lipid layers in combination with molecular adaptors to selectively adhere the viruses to the AG surface. This further development of the AG method may prove essential for the gentle and selective purification of enveloped viruses directly onto EM grids for ultrastructural analyses.

scholarly journals Site Specific Three-dimensional Structural Analysis in Tissues and Cells Using Automated DualBeam Slice &View

2007 ◽  
Vol 15 (2) ◽  
pp. 26-31 ◽  
Author(s):  
Ben Lich
Keyword(s):  
Electron Microscopy ◽  
Focused Ion Beam ◽  
Ion Beam ◽  
Three Dimensional ◽  
Surface Region ◽  
Dimensional Structure ◽  
Near Surface ◽  
Imaging Capability ◽  
Fluorescent Markers ◽  
Confocal And Electron Microscopy

DualBeam instruments that combine the imaging capability of scanning electron microscopy (SEM) with the cutting and deposition capability of a focused ion beam (FIB) provide biologists with a powerful tool for investigating three-dimensional structure with nanoscale (1 nm-100 nm) resolution. Ever since Van Leeuwenhoek used the first microscope to describe bacteria more than 300 years ago, microscopy has played a central role in scientists' efforts to understand biological systems. Light microscopy is generally limited to a useful resolution of about a micrometer. More recently the use of confocal and electron microscopy has enabled investigations at higher resolution. Used with fluorescent markers, confocal microscopy can detect and localize molecular scale features, but its imaging resolution is still limited. SEM is capable of nanometer resolution, but is limited to the near surface region of the sample.

Supercoils in plant DNA: nucleoid sedimentation studies

1988 ◽  
Vol 89 (2) ◽  
pp. 243-252
Author(s):  
L.M. Stoilov ◽  
J.S. Zlatanova ◽  
A.P. Vassileva ◽  
M.G. Ivanchenko ◽  
C.P. Krachmarov ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Gamma Irradiation ◽  
Protein Content ◽  
Ethidium Bromide ◽  
Three Dimensional ◽  
The Other ◽  
Dimensional Structure ◽  
Salt Treatment ◽  
Plant Dna ◽  
Ionic Detergent

Plant nuclei have been studied with respect to the three-dimensional structure of DNA. Nucleoids derived from nuclei by non-ionic detergent and high salt treatment were analysed by sedimentation in a series of sucrose gradients containing increasing amounts of the intercalating agent ethidium bromide. In addition the nucleoid sedimentation behaviour was investigated following gamma irradiation. The results show that plant DNA is supercoiled, as is the DNA from the other eukaryotes studied, and contains approximately the same concentration of superhelical turns but probably relatively fewer DNA superhelical loops. The plant nuclear populations in all cases studied give rise to two distinct nucleoid bands. These have been characterized by electron microscopy and by their DNA and protein content. The possible origin of the two bands is discussed.

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