In Situ Hybridization tor RNA: Nonradioactive Probe: Oligo-DNA Probe: Digoxigenin (II)

2000 ◽  
pp. 182-195
Author(s):  
Masanobu Miyazaki ◽  
Yoshiyuki Ozono ◽  
Takashi Harada ◽  
Shigeru Kohno
Keyword(s):  
In Situ Hybridization ◽  
Dna Probe

Ultrastructural analysis of the spatial distribution of MRNA

1992 ◽  
Vol 50 (1) ◽  
pp. 552-553
Author(s):  
Gary Bassell ◽  
Robert H. Singer
Keyword(s):  
Electron Microscopy ◽  
Spatial Distribution ◽  
In Situ Hybridization ◽  
High Resolution ◽  
Nucleic Acids ◽  
Colloidal Gold ◽  
Dna Probe ◽  
Nucleotide Analogs ◽  
Gold Particles

We have been investigating the spatial distribution of nucleic acids intracellularly using in situ hybridization. The use of non-isotopic nucleotide analogs incorporated into the DNA probe allows the detection of the probe at its site of hybridization within the cell. This approach therefore is compatible with the high resolution available by electron microscopy. Biotinated or digoxigenated probe can be detected by antibodies conjugated to colloidal gold. Because mRNA serves as a template for the probe fragments, the colloidal gold particles are detected as arrays which allow it to be unequivocally distinguished from background.

In Situ Hybridization at the Electron Microscopic Level Using a Bromodeoxyuridine Labeled DNA Probe

1993 ◽  
Vol 68 (3) ◽  
pp. 169-174 ◽  
Author(s):  
Chong-Hua Yao ◽  
Sohei Kitazawa ◽  
Takahiro Fujimori ◽  
Sakan Maeda
Keyword(s):  
In Situ Hybridization ◽  
Microscopic Level ◽  
Dna Probe ◽  
Electron Microscopic Level ◽  
Electron Microscopic

Detection of an i(17q) chromosome by fluorescent in situ hybridization with a chromosome 17 alpha satellite DNA probe

1992 ◽  
Vol 62 (2) ◽  
pp. 140-143 ◽  
Author(s):  
Hitoshi Nakagawa ◽  
Johji Inazawa ◽  
Shinichi Misawa ◽  
Shinji Tanaka ◽  
Teruyuki Takashima ◽  
...  
Keyword(s):  
In Situ Hybridization ◽  
Satellite Dna ◽  
Fluorescent In Situ Hybridization ◽  
Dna Probe ◽  
Chromosome 17 ◽  
Alpha Satellite ◽  
Alpha Satellite Dna

scholarly journals The effect of avidin-biotin interactions in detection systems for in situ hybridization.

1992 ◽  
Vol 40 (1) ◽  
pp. 135-141 ◽  
Author(s):  
E J Speel ◽  
B Schutte ◽  
F C Ramaekers ◽  
A H Hopman
Keyword(s):  
In Situ Hybridization ◽  
Network Formation ◽  
Transitional Cell ◽  
Fluorescein Isothiocyanate ◽  
Staining Technique ◽  
Dna Probe ◽  
Detection Systems ◽  
Amplification Method ◽  
The Individual

The effect of avidin-biotin interactions in several detection systems for the non-radioactive in situ hybridization (ISH) technique was studied in a model system using a transitional cell carcinoma line and a biotinylated DNA probe. We performed fluorescence ISH to unravel the individual steps in a sensitive and frequently used amplification method which makes use of the alternating cytochemical detection layers of fluorescein isothiocyanate-conjugated avidin (AvFITC) and biotinylated goat anti-avidin (BioGAA) antibodies to detect the hybridized and biotinylated probe. Our experiments revealed that BioGAA antibodies bind with their antigen binding sites and not with their biotin moieties to avidin molecules that have already interacted with the DNA probe. The probable working mechanism of this amplification method is presented in a model. Furthermore, we used a peroxidase staining technique to compare with each other the sensitivity of several other detection systems in which avidin-biotin interactions play an important role, e.g., the avidin-biotinylated peroxidase complex (ABC) system. The experiments show that avidin molecules can not be efficiently used to interconnect two biotinylated molecular layers, since their introduction leads to firmly closed cytochemical networks. Such a closed network is already formed between the hybridized and biotinylated DNA probe and a first detection layer of avidin molecules, as appears from the finding that biotinylated molecules could hardly be coupled to these avidin molecules in a following detection layer. Therefore, the results presented here provide us with new insight into the molecular basis of cytochemical network formation. This will enable us to choose the proper procedures for increasing the sensitivity of ISH detection systems.

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