Preparation of plant cells for transmission electron microscopy to optimize immunogold labeling of carbohydrate and protein epitopes

2012 ◽  
Vol 7 (9) ◽  
pp. 1716-1727 ◽  
Author(s):  
Sarah M Wilson ◽  
Antony Bacic
Keyword(s):  
Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Plant Cells ◽  
Immunogold Labeling ◽  
Transmission Electron

Ultrastructural Morphology of the Plastids in Ferocactus Latispinus (Haworth)

1981 ◽  
Vol 39 ◽  
pp. 648-649
Author(s):  
E. R. Rivera
Keyword(s):  
Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Apical Meristem ◽  
Plant Cells ◽  
Cortical Cells ◽  
Conventional Transmission Electron Microscopy ◽  
Meristematic Cells ◽  
Dense Cytoplasm ◽  
Transmission Electron ◽  
Peripheral Cytoplasm

Mature plants of Ferocactus latispinus were divided into the apical meristem, cortical and cambial tissue, and young and mature photosynthetic tissue. Pieces 2x2x4 mm were fixed in 0.075 M PIPES buffered 5% glutaraldehyde and further treated for conventional transmission electron microscopy.The shoot meristematic cells contained dense cytoplasm with a few small vacuoles. All organelles expected to be found in unspecialized plant cells were present. The proplastids were 1-2 diameters larger than mitochondria. Some of these plastids were elongated and most contained small amounts of lipid globules in the stroma (Fig. 1). In addition, phytoferritin was sometimes found in the stroma. No starch was found in these organelles. The thylakoids were poorly developed, and when there were no inclusions in the stroma, proplastids were difficult to distinguish from mitochondria (Fig. 1).Cortical cells had very large, well-developed vacuoles. The organelles were restricted to the peripheral cytoplasm of the cell and to a few transvacuolar cytoplasmic strands.

Immunogold labeling of HTLV-III/LAV in H9 cells studied by transmission electron microscopy

1986 ◽  
Vol 13 (3) ◽  
pp. 265-269 ◽  
Author(s):  
D. Pekovic ◽  
S. Garzon ◽  
H. Strykowski ◽  
D. Ajdukovic ◽  
M. Gornitsky ◽  
...  
Keyword(s):  
Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Immunogold Labeling ◽  
Transmission Electron

scholarly journals Pitfalls of immunogold labeling: analysis by light microscopy, transmission electron microscopy, and photoelectron microscopy.

1987 ◽  
Vol 35 (8) ◽  
pp. 843-853 ◽  
Author(s):  
G B Birrell ◽  
K K Hedberg ◽  
O H Griffith
Keyword(s):  
Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Light Microscopy ◽  
Tannic Acid ◽  
Cultured Cells ◽  
Immunogold Labeling ◽  
Fish Gelatin ◽  
Nonspecific Binding ◽  
Gold Particles ◽  
Transmission Electron

The immunogold method is widely used to localize, identify, and distinguish cellular antigens. There are, however, some pitfalls that can lead to nonspecific binding, particularly in cytoskeletal studies with gold probes prepared from small gold particles. We present a list of suggestions for minimizing nonspecific binding, with particular attention to two problems identified in this study. First, we find that the method used to prepare the colloidal gold particles affects the degree of nonspecific binding. Second, the standard BSA-stabilized small gold probes evidently possess exposed regions that bind to the proteins of cytoskeletal preparations. This was investigated in whole-mount cytoskeletal preparations of cultured cells by use of light microscopy, transmission electron microscopy, and photoelectron microscopy of silver-enhanced specimens. Gold probes were made from approximately 5-nm particles generated by reduction of HAuCl4 with three different reducing agents: white phosphorus, sodium borohydride, and citrate-tannic acid. All three preparations stabilized in the conventional way showed significant levels of nonspecific binding, which was highest with citrate-tannic acid. This problem was largely solved with all three types of probes by including fish gelatin in the probe buffer, by substituting fish gelatin for the BSA stabilizer used to prepare the probes, or by pre-adsorption methods. Application of these techniques resulted in clear immunogold labeling patterns with minimal nonspecific background.

Ultrastructure and immunogold labeling of cytoplasmic fibrous bundles in tobacco leaf cells infected with tobacco vein mottling virus

1992 ◽  
Vol 50 (1) ◽  
pp. 676-677
Author(s):  
E. D. Ammar ◽  
D. W. Thombury ◽  
T. P. Pirone
Keyword(s):  
Electron Microscopy ◽  
Transmission Electron Microscopy ◽  
Immunogold Labeling ◽  
Infected Leaf ◽  
Transmission Electron ◽  
Aphid Vectors ◽  
Cylindrical Inclusions ◽  
Tobacco Vein Mottling Virus ◽  
Unknown Composition ◽  
Infected Tobacco

Tobacco vein mottling virus (TVMV) is a potyvirus, with flexuous filamentous particles ca. 765 nm long and 12-13 nm in diameter. TVMV is transmitted by sap inoculation and by aphid vectors. Cylindrical inclusions characteristic of infections with potyviruses have been found in the cytoplasm of TVMV-infected tobacco leaves and protoplasts, and the presence of fibrous inclusions of unknown composition in the cytoplasm and nuclei of TVMV-infected tobacco has been reported. Here, we report the results of transmission electron microscopy (TEM) and immunogold labeling of cytoplasmic fibrous bundles found in TVMV-infected leaf cells, providing evidence that these bundles are composed of TVMV virions and/or coat protein.

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