Journal of Histochemistry and Cytochemistry Priciples for Free Access to Science
  Search:   
    >> Advanced Search

Guidelines | Subscriptions | About | exPRESS - Current - Archive | Business Information | Contact
This Article
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Similar articles in this journal
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Right arrow reprints & permissions
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Speel, E. J.
Right arrow Articles by Hopman, A. H.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Speel, E. J.
Right arrow Articles by Hopman, A. H.
Social Bookmarking
Add to CiteULike   Add to Complore   Add to Connotea   Add to Del.icio.us   Add to Digg   Add to Reddit   Add to Technorati  
What's this?

A novel fluorescence detection method for in situ hybridization, based on the alkaline phosphatase-fast red reaction

EJ Speel, B Schutte, J Wiegant, FC Ramaekers and AH Hopman

Department of Molecular Cell Biology and Genetics, University of Limburg, Maastricht, The Netherlands.

We have used naphthol-ASMX-phosphate and Fast Red TR in combination with alkaline phosphatase (APase) to produce fluorescent precipitated reaction products in a non-radioactive in situ hybridization (ISH) method. To obtain optimal and discrete localization of the strongly red fluorescent ISH signals, the enzyme precipitation procedure was optimized. The optimal reaction time and the concentrations of substrate and capture agent were determined. Furthermore, polyvinyl alcohol (PVA) was used to increase the viscosity of the reaction mixture and thus to reduce diffusion of the reaction product. Our results show that the APase-Fast Red detection method has at least the same sensitivity as currently observed in other immunofluorescent detection systems. A single copy DNA sequence of 15.8 KB could be localized with high efficiency in metaphase spreads and in interphase nuclei. Double labeling procedures, in which the FITC- and azo-dye fluorescence are combined, are also feasible. The red fluorescent ISH signals showed hardly any fading as compared with FITC fluorescence on exposure to either light from the mercury-arc lamp or laser light. Therefore, these red fluorescent signals with a virtually permanent character allow a better analysis and three-dimensional localization of such cytochemically detected genomic fractions by means of confocal scanning laser microscopy as compared with the use of FITC, TRITC, or Texas Red as label.

Volume 40, Issue 9, pp. 1299-1308, 09/01/1992
Copyright © 1992 by The Histochemical Society


Add to CiteULike CiteULike   Add to Complore Complore   Add to Connotea Connotea   Add to Del.icio.us Del.icio.us   Add to Digg Digg   Add to Reddit Reddit   Add to Technorati Technorati    What's this?


This article has been cited by other articles:


Home page
 J. Gen. Virol.Home page
K. W. R. van Cleef, W. M. A. Scaf, K. Maes, S. J. F. Kaptein, E. Beuken, P. S. Beisser, F. R. M. Stassen, G. E. L. M. Grauls, C. A. Bruggeman, and C. Vink
The rat cytomegalovirus homologue of parvoviral rep genes, r127, encodes a nuclear protein with single- and double-stranded DNA-binding activity that is dispensable for virus replication
J. Gen. Virol., July 1, 2004; 85(7): 2001 - 2013.
[Abstract] [Full Text] [PDF]

Home page
 J. Histochem. Cytochem.Home page
A. N. Player, L.-P. Shen, D. Kenny, V. P. Antao, and J. A. Kolberg
Single-copy Gene Detection Using Branched DNA (bDNA) In Situ Hybridization
J. Histochem. Cytochem., May 1, 2001; 49(5): 603 - 612.
[Abstract] [Full Text]

Home page
 Am. J. Pathol.Home page
L. Ermert, A. C. Hocke, H.-R. Duncker, W. Seeger, and M. Ermert
Comparison of Different Detection Methods in Quantitative Microdensitometry
Am. J. Pathol., February 1, 2001; 158(2): 407 - 417.
[Abstract] [Full Text] [PDF]

Home page
 J. Histochem. Cytochem.Home page
J. F. Breininger and D. G. Baskin
Fluorescence In Situ Hybridization of Scarce Leptin Receptor mRNA using the Enzyme-Labeled Fluorescent Substrate Method and Tyramide Signal Amplification
J. Histochem. Cytochem., December 1, 2000; 48(12): 1593 - 1600.
[Abstract] [Full Text]

Home page
 J. Histochem. Cytochem.Home page
M. Grino and A. J. Zamora
An In Situ Hybridization Histochemistry Technique Allowing Simultaneous Visualization by the Use of Confocal Microscopy of Three Cellular mRNA Species in Individual Neurons
J. Histochem. Cytochem., June 1, 1998; 46(6): 753 - 760.
[Abstract] [Full Text]

Home page
 J. Histochem. Cytochem.Home page
E. J.M. Speel, F. C.S. Ramaekers, and A. H.N. Hopman
Sensitive Multicolor Fluorescence In Situ Hybridization Using Catalyzed Reporter Deposition (CARD) Amplification
J. Histochem. Cytochem., October 1, 1997; 45(10): 1439 - 1446.
[Abstract] [Full Text] [PDF]

Home page
 J. Histochem. Cytochem.Home page
V. B. Paragas, Y.-Z. Zhang, R. P. Haugland, and V. L. Singer
The ELF-97 Alkaline Phosphatase Substrate Provides a Bright, Photostable, Fluorescent Signal Amplification Method for FISH
J. Histochem. Cytochem., March 1, 1997; 45(3): 345 - 358.
[Abstract] [Full Text] [PDF]

Home page
 J. Histochem. Cytochem.Home page
R. P. M. van Gijlswijk, H. J. M. A. A. Zijlmans, J. Wiegant, M. N. Bobrow, T. J. Erickson, K. E. Adler, H. J. Tanke, and A. K. Raap
Fluorochrome-labeled Tyramides: Use in Immunocytochemistry and Fluorescence In Situ Hybridization
J. Histochem. Cytochem., March 1, 1997; 45(3): 375 - 382.
[Abstract] [Full Text] [PDF]



Guidelines | Subscriptions | About | exPRESS - Current - Archive | Business Information | Contact
The Journal of Histochemistry & Cytochemistry is owned, published, and licensed by The Histochemical Society © 1992