Confocal laser microscopy of impregnated central nervous system tissue

Author(s):  
J.N. Turner ◽  
M. Siemens ◽  
D. Szarowski ◽  
D.N. Collins

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).

2005 ◽  
Vol 68 (3) ◽  
pp. 630-632 ◽  
Author(s):  
EVA RENCOVA

An indirect competitive enzyme-linked immunosorbent assay was used to compare four commercially available monoclonal and polyclonal antibodies prepared against the markers of central nervous system tissue: glial fibrillary acidic protein, neurofilaments, and myelin basic protein. These antibodies were investigated for their usefulness at the detection of prohibited central nervous system tissue in meat products to prevent spread of bovine spongiform encephalopathy and its human form, a new variant of Creutzfeldt-Jakob disease. The detection limit was 2.5 to 10%. The specificity was tested using extracts of tissue samples of bovine and porcine brain, spinal cord, lymphatic node, diaphragm, skeletal muscle, and peripheral nerves. All antibodies reacted with both bovine and porcine tissue extracts.


Author(s):  
T.D. Pollard ◽  
P. Maupin

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.


Author(s):  
G. E. Tyson ◽  
M. J. Song

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.


2000 ◽  
Vol 79 (8) ◽  
pp. 635-639 ◽  
Author(s):  
TOSHIYUKI HATA ◽  
TOSHIHIRO YANAGIHARA ◽  
MINAKO MATSUMOTO ◽  
UIKO HANAOKA ◽  
MARI UETA ◽  
...  

2013 ◽  
Vol 20 (1) ◽  
pp. 164-174 ◽  
Author(s):  
Gabriella Kiss ◽  
Xuemin Chen ◽  
Melinda A. Brindley ◽  
Patricia Campbell ◽  
Claudio L. Afonso ◽  
...  

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.


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