Gold Cluster Compounds are Useful Immunoprobes

2000 ◽  
Vol 6 (S2) ◽  
pp. 328-329
Author(s):  
J.M. Robinson ◽  
T. Takizawa ◽  
D.D. Vandré
Keyword(s):  
Electron Microscope ◽  
Gold Cluster ◽  
Cluster Compounds ◽  
Microscope Level ◽  
Reporter System ◽  
Electron Microscope Level ◽  
Light Microscope Level ◽  
Antibody Recognition ◽  
Particulate Probes

Immunocytochemistry generally refers to methods directed toward obtaining information on the in situ distribution of antigens in cells and tissues. Immunocytochemical methods can be applied at either the light or electron microscope levels, or both in concert. The detection of antibody recognition of an antigen (i.e., localization of an antigen) relies upon a reporter system. At the light microscope level, enzymes (e.g., horseradish peroxidase) or fluorochromes are the most widely used reporters in immunocytochemistry. At the electron microscope level, particulate probes (e.g., colloidal gold) are the most widely used reporters. However, enzymes and even fluorochromes can be used at the EM level.In this abstract, we discuss our use of gold cluster immunoprobes as the reporter system in both light and electron microscope level immuncytochemistry. These gold cluster immunoprobes, are commercially known as NanogoldTM (NG). These probes are very small with a diameter of 1.4-nm.

Molecular hybridization of RNA and DNA in situ: visualization at the electron microscope level

1976 ◽  
Vol 106 (2) ◽  
pp. 185-198 ◽  
Author(s):  
J. Jacob ◽  
M. H. Moar ◽  
Katherine Gillies ◽  
D. Macleod ◽  
K. W. Jones
Keyword(s):  
Electron Microscope ◽  
Molecular Hybridization ◽  
Microscope Level ◽  
Electron Microscope Level ◽  
In Situ Visualization ◽  
Rna And Dna

scholarly journals In situ hybridization at the electron microscope level: hybrid detection by autoradiography and colloidal gold

1982 ◽  
Vol 95 (2) ◽  
pp. 609-618 ◽  
Author(s):  
NJ Hutchison ◽  
PR Langer-Safer ◽  
DC Ward ◽  
BA Hamkalo
Keyword(s):  
In Situ Hybridization ◽  
Electron Microscope ◽  
Satellite Dna ◽  
Light Microscope ◽  
Test System ◽  
Microscope Level ◽  
Metaphase Chromosomes ◽  
Light Microscope Level ◽  
Mouse Satellite Dna

In situ hybridization has become a standard method for localizing DNA or RNA sequences in cytological preparations. We developed two methods to extend this technique to the transmission electron microscope level using mouse satellite DNA hybridization to whole mount metaphase chromosomes as the test system. The first method devised is a direct extension of standard light microscope level using mouse satellite DNA hybridization to whole mount metaphase chromosomes as the test system. The first method devised is a direct extension of standard light microscope in situ hybridization. Radioactively labeled complementary RNA (cRNA) is hybridized to metaphase chromosomes deposited on electron microscope grids and fixed in 70 percent ethanol vapor; hybridixation site are detected by autoradiography. Specific and intense labeling of chromosomal centromeric regions is observed even after relatively short exposure times. Inerphase nuclei present in some of the metaphase chromosome preparations also show defined paatterms of satellite DNA labeling which suggests that satellite-containing regions are associate with each other during interphase. The sensitivity of this method is estimated to at least as good as that at the light microscope level while the resolution is improved at least threefold. The second method, which circumvents the use of autoradiogrphic detection, uses biotin-labeled polynucleotide probes. After hybridization of these probes, either DNA or RNA, to fixed chromosomes on grids, hybrids are detected via reaction is improved at least threefold. The second method, which circumvents the use of autoradiographic detection, uses biotin-labeled polynucleotide probes. After hybridization of these probes, either DNA or RNA, to fixed chromosomes on grids, hybrids are detected via reaction with an antibody against biotin and secondary antibody adsorbed to the surface of over centromeric heterochromatin and along the associated peripheral fibers. Labeling is on average ten times that of background binding. This method is rapid and possesses the potential to allow precise ultrastructual localization of DNA sequences in chromosomes and chromatin.

scholarly journals In situ hybridization at the electron microscope level: localization of transcripts on ultrathin sections of Lowicryl K4M-embedded tissue using biotinylated probes and protein A-gold complexes.

1986 ◽  
Vol 102 (5) ◽  
pp. 1646-1653 ◽  
Author(s):  
M Binder ◽  
S Tourmente ◽  
J Roth ◽  
M Renaud ◽  
W J Gehring
Keyword(s):  
Electron Microscope ◽  
Protein A ◽  
Microscope Level ◽  
Gold Complex ◽  
Electron Microscope Level ◽  
Thin Sections ◽  
Hybrid Detection ◽  
Ultrathin Sections ◽  
Lowicryl K4m

A technique has been developed for localizing hybrids formed in situ on semi-thin and ultrathin sections of Lowicryl K4M-embedded tissue. Biotinylated dUTP (Bio-11-dUTP and/or Bio-16-dUTP) was incorporated into mitochondrial rDNA and small nuclear U1 probes by nick-translation. The probes were hybridized to sections of Drosophila ovaries and subsequently detected with an anti-biotin antibody and protein A-gold complex. On semi-thin sections, probe detection was achieved by amplification steps with anti-protein A antibody and protein A-gold with subsequent silver enhancement. At the electron microscope level, specific labeling was obtained over structures known to be the site of expression of the appropriate genes (i.e., either over mitochondria or over nuclei). The labeling pattern at the light microscope level (semi-thin sections) was consistent with that obtained at the electron microscope level. The described nonradioactive procedures for hybrid detection on Lowicryl K4M-embedded tissue sections offer several advantages: rapid signal detection: superior morphological preservation and spatial resolution; and signal-to-noise ratios equivalent to radiolabeling.

Genomic in situ hybridization to sectioned nuclei shows chromosome domains in grass hybrids

1990 ◽  
Vol 95 (3) ◽  
pp. 335-341
Author(s):  
A.R. Leitch ◽  
W. Mosgoller ◽  
T. Schwarzacher ◽  
M.D. Bennett ◽  
J.S. Heslop-Harrison
Keyword(s):  
In Situ Hybridization ◽  
Genomic Dna ◽  
Microscope Level ◽  
Hordeum Chilense ◽  
Root Tip ◽  
Interphase Nuclei ◽  
Light Microscope Level ◽  
Thin Sections ◽  
Electron Microscopes

In situ hybridization using biotinylated total genomic DNA and avidin detection systems was adapted for examination of thin-sectioned plant material in the light and electron microscopes. Root tip material was preserved prior to sectioning, so that the in vivo disposition of the chromatin was maintained. Use of total genomic DNA from Secale africanum as a probe enabled the chromatin from the two parental genomes in the grass hybrid Hordeum chilense × S. africanum to be distinguished. The biotinylated probe preferentially labelled the chromosomes of S. africanum origin. DNA-DNA hybrids were visualized at the light-microscope level by Texas Red fluorescence and at the electron-microscope level by the enzymic precipitation of DAB (diaminobenzidine) or by colloidal gold particles. The use of thin sections allowed the location of probe hybridization to be established unequivocally in both metaphase and interphase nuclei. Analysis of interphase nuclei showed that chromatin originating from the two parental genomes did not intermix but occupied distinct domains.

Sequential Changes Associated with Cell Wall Formation and Fusion in the Vascular Cambium

IAWA Journal ◽  
1981 ◽  
Vol 2 (4) ◽  
pp. 151-162 ◽  
Author(s):  
A.M. Catesson ◽  
J.C. Roland
Keyword(s):  
Cell Wall ◽  
Electron Microscope ◽  
Cell Walls ◽  
Microscope Level ◽  
Cell Wall Formation ◽  
Characteristic Sequence ◽  
Electron Microscope Level ◽  
Starting Point ◽  
Cell Wall Development ◽  
Cambial Zone

Cytochemical techniques and mild extractions were used at the electron microscope level for the study of the cambial zone of several hardwoods and one softwood. The maturation processes of the primary radial and tangential cell walls involve a progressive disappearance of their initial heterogeneity. The buttress-like zone joining these walls appears to be the starting point for a characteristic sequence of changes and intra-wall rearrangement. Topochemical results have suggested an alternative to the 'emboxing concept' of cell wall development.

scholarly journals CYTOCHEMISTRY OF 5-HYDROXYTRYPTAMINE AT THE ELECTRON MICROSCOPE LEVEL I. STUDY OF THE SPECIFICITY OF THE REACTION IN ISOLATED BLOOD PLATELETS

1968 ◽  
Vol 16 (3) ◽  
pp. 162-171 ◽  
Author(s):  
GUILLERMO JAIM ETCHEVERRY ◽  
LUIS MARIA ZIEHER
Keyword(s):  
Electron Microscope ◽  
Blood Platelets ◽  
Microscope Level ◽  
Storage Site ◽  
Electron Microscope Level ◽  
Dense Granules ◽  
Amine Content ◽  
Level I ◽  
Previous Assumption

Blood platelets obtained from normal rabbits and those isolated from reserpine-treated animals and subsequently incubated in vitro with 5-hydroxytryptamine, norepinephrine and histamine were assayed for amine content or processed for examination under the electron microscope. With the glutaraldehyde-dichromate reaction for unsubstituted catechol- and indoleamines, reactive granules were observed in normal platelets. Formaldehyde fixation prior to the glutaraldehyde-dichromate reaction resulted in a similar image under the electron microscope. In platelets obtained from animals treated with reserpine a decrease of the amine content with a corresponding reduction in the number of dense granules was observed. Following incubation with 5-hydroxytryptamine the concentration of the amine increased markedly and the number of dense granules that reacted with both techniques became practically normal. In norepinephrine-incubated platelets dense granules were demonstrated with the glutaraldehyde-dichromate reaction, but no reactive products were observed using prefixation with formaldehyde. Histamine was also incorporated into depleted platelets but gave no reaction. It is concluded that prefixation with formaldehyde renders negative the reaction with catecholamines, leaving unaffected indoleamine-reactive sites. The previous assumption that the dense granules contain 5-hydroxytryptamine has been confirmed by such a cytochemical approach. The possibility that these organelles constitute a common storage site for different amines is discussed.

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