In situ localization of chalcone synthase inLarix needles by indirect immunofluorescence

PROTOPLASMA ◽  
1989 ◽  
Vol 153 (1-2) ◽  
pp. 58-61 ◽  
Author(s):  
A. Lembach ◽  
L. Beerhues ◽  
R. Wiermann
Keyword(s):  
Indirect Immunofluorescence ◽  
Chalcone Synthase

Multiple Iteractive Labeling by Antibody Neodeposition (MILAN)

2019 ◽  
Author(s):  
Giorgio Cattoretti ◽  
Francesca Maria Bosisio ◽  
Lukas Marcelis ◽  
Maddalena Maria Bolognesi
Keyword(s):  
Image Analysis ◽  
Tissue Section ◽  
Indirect Immunofluorescence ◽  
Analysis Software ◽  
Tissue Sections ◽  
Image Analysis Software ◽  
Single Section ◽  
Fluorescent Image ◽  
Secondary Antibodies

Abstract Multiplexing, labeling for multiple immunostains the very same cell or tissue section in situ, is of considerable interest. The major obstacles to the diffusion of this technique are high costs in custom antibodies and instruments, low throughput, scarcity of specialized skills or facilities. We have validated and detail here a method based on common primary and secondary antibodies, diffusely available fluorescent image scanners and routinely processed tissue sections \(FFPE). It entails rounds of four-color indirect immunofluorescence, image acquisition and removal \(stripping) of the antibodies, before another stain is applied. The images are digitally registered and the autofluorescence is subtracted. Removal of antibodies is accomplished by disulphide cleavage. In excess of 50 different antibody stains can be applied to one single section from routinely fixed and embedded tissue. This method requires a modest investment in hardware and materials and uses freeware image analysis software.

Chemical components and their locations in the Verticillium fungicola cell wall

2000 ◽  
Vol 46 (2) ◽  
pp. 101-109 ◽  
Author(s):  
M Calonje ◽  
M Novaes-Ledieu ◽  
D Bernardo ◽  
O Ahrazem ◽  
C García Mendoza
Keyword(s):  
Indirect Immunofluorescence ◽  
Cell Walls ◽  
Agaricus Bisporus ◽  
Chemical Structure ◽  
Wall Material ◽  
Chemical Components ◽  
Carbohydrate Polymers ◽  
Verticillium Fungicola ◽  
Intermediate Layers

The chemical structure of cell walls and fractions of Verticillium fungicola, a pathogen of Agaricus bisporus, as well as their corresponding ultrastructures were studied. There are at least three chemically distinct types of carbohydrate polymers: one yielding mannose with lower amounts of galactose and glucose (glucogalactomannan), another one composed mainly of glucose (glucan), and a third one containing only N-acetylglucosamine (chitin). Attempts were made to locate these materials in situ by comparing electron micrographs of shadowed and sectioned cell walls, and also by indirect immunofluorescence. It was shown that none of these polymers constituted a completely physically distinct layer, but there seem to be different solubility properties in the outer, inner, and intermediate layers. It was also shown that fibrillar material (chitin) embedded in cementing glucan constituted the residual inner fraction of the original wall material. Indirect immunofluorescence showed the location of a significant amount of glucogalactomannan on the surface of the walls in which rodlet structures were visualized by electron microscopy.Key words: cell walls, polysaccharides, Verticillium fungicola.

scholarly journals Simple diagnostic test for antibodies to Encephalitozoon cuniculi based on enzyme immunoassay

1981 ◽  
Vol 15 (1) ◽  
pp. 41-44 ◽  
Author(s):  
J. C. Cox ◽  
R. Horsburgh ◽  
D. Pye
Keyword(s):  
Cell Culture ◽  
Enzyme Immunoassay ◽  
Diagnostic Test ◽  
Indirect Immunofluorescence ◽  
Antigen Processing ◽  
Encephalitozoon Cuniculi ◽  
Immunofluorescence Test ◽  
Indirect Immunofluorescence Test ◽  
Specialized Equipment

Rabbit antibodies against Encephalitozoon cuniculi were detected in an enzyme immunoassay procedure in which antigen was grown and used in situ. The test appeared to be more sensitive than the indirect immunofluorescence test with which it was compared, but gave essentially the same results for the 64 sera evaluated. This procedure will allow any laboratory with cell-culture facilities to produce a diagnostic antigen without the need for antigen processing. It is simple and reliable, and does not require specialized equipment or microscopic assessment.

scholarly journals Detection of Streptococcus Suis by in Situ Hybridization, Indirect Immunofluorescence, and Peroxidase-Antiperoxidase Assays in Formalin-Fixed, Paraffin-Embedded Tissue Sections from Pigs

2000 ◽  
Vol 12 (3) ◽  
pp. 224-232 ◽  
Author(s):  
Mette Boye ◽  
Anne A. Feenstra ◽  
Conny Tegtmeier ◽  
Lars Ole Andresen ◽  
Søren R. Rasmussen ◽  
...  
Keyword(s):  
In Situ Hybridization ◽  
Indirect Immunofluorescence ◽  
Polyclonal Antibodies ◽  
Single Cells ◽  
Streptococcus Suis ◽  
Tissue Sections ◽  
Formalin Fixed Paraffin ◽  
Formalin Fixed Paraffin Embedded ◽  
Formalin Fixed

Streptococcus suis is an important pathogen in pigs and is considered a zoonotic agent. To aid diagnosis of infection caused by S. suis, a species-specific probe targeting 16S ribosomal RNA was designed and used for fluorescent in situ hybridization. Two additional immunohistochemical detection methods, an indirect immunofluorescence assay and a peroxidase-antiperoxidase method, using polyclonal antibodies also were developed. The specificity of the oligonucleotide probe was examined by whole-cell and dot-blot hybridization against reference strains of the 35 serotypes of S. suis and other closely related streptococci and other bacteria commonly isolated from pigs. The probe was specific for S. suis serotypes 1–31. The specificity of the polyclonal antibodies, which has previously been evaluated for use in diagnostic bacteriology for typing of serotype 2, was further evaluated in experimentally infected murine tissue with pure culture of different serotypes of S. suis, related streptococci, and other bacteria commonly found in pigs. The polyclonal antibodies against S. suis serotype 2 cross-reacted with serotypes 1 and 1/2 in these assays. The in situ hybridization and the immunohistochemical methods were used for detection of S. suis in formalin-fixed, paraffin-embedded tissue sections of brain, endocardium, and lung from pigs infected with S. suis. The methods developed were able to detect single cells of S. suis in situ in the respective samples, whereas no signal was observed from control tissue sections that contained organisms other than S. suis. These techniques are suitable for determining the in vivo localization of S. suis for research and diagnostic purposes.

scholarly journals In situ localization of the estrogen receptor in living cells with the fluorescent phytoestrogen coumestrol.

1993 ◽  
Vol 41 (6) ◽  
pp. 801-810 ◽  
Author(s):  
R J Miksicek
Keyword(s):  
Estrogen Receptor ◽  
Indirect Immunofluorescence ◽  
Cultured Cells ◽  
Living Cells ◽  
Receptor Protein ◽  
Intact Cells ◽  
Naturally Occurring ◽  
Efficient Detection ◽  
Human Estrogen Receptor

Coumestrol is a naturally occurring plant coumarin that displays high affinity for the hormone-binding site of the human estrogen receptor (hER), for which it serves as a potent non-steroidal agonist. Coumestrol emits intense blue fluorescence when bound to this protein, making it ideally suited for use as a cytological stain to detect ER in fixed and intact cells. Conditions are reported for the efficient detection of recombinant ER protein artificially expressed in cultured cells by calcium phosphate-mediated transfection. Coumestrol fluorescence co-localizes with hER protein detected by indirect immunofluorescence with an hER-specific anti-peptide antiserum, confirming the specificity of this reagent. Fluorescence from ER occupied by coumestrol shows a predominantly nuclear distribution, even when the receptor protein is cross-linked in situ by fixation with glutaraldehyde plus paraformaldehyde before coumestrol exposure. This corroborates previous conclusions, based on indirect immunofluorescence analysis, that the ER remains nuclear in the absence of hormone. Examination of the pattern of coumestrol staining in mitotic cells further indicates that hER remains associated with condensed chromatin. Such observations illustrate the potential for using coumestrol to investigate real-time effects of a variety of physiological stimuli on the subcellular distribution of hER in living cells.

In situ localization of chalcone synthase in tannin-containing plants

1993 ◽  
Vol 32 (3) ◽  
pp. 585-590 ◽  
Author(s):  
B. Karwatzki ◽  
A. Herget ◽  
L. Beerhues ◽  
R. Wiermann
Keyword(s):  
Chalcone Synthase

scholarly journals In-situ localization of chalcone synthase mRNA in pea root nodule development

1992 ◽  
Vol 2 (2) ◽  
pp. 143-151 ◽  
Author(s):  
Wei-Cai Yang ◽  
Hayo C.J. Canter Cramers ◽  
Peter Hogendijk ◽  
Panagiotis Katinakis ◽  
Carel A. Wijffelman ◽  
...  
Keyword(s):  
Root Nodule ◽  
Chalcone Synthase ◽  
Nodule Development ◽  
Pea Root
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