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
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 Nuovo, G. J.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Nuovo, G. J.
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?
Journal of Histochemistry and Cytochemistry, Vol. 49, 1329-1340, November 2001, Copyright © 2001, The Histochemical Society, Inc.

REVIEW

Co-labeling Using In Situ PCR: A Review

Gerard J. Nuovoa
a Department of Pathology, Ohio State University Medical Center, Columbus, Ohio

Correspondence to: Gerard J. Nuovo, Dept. of Pathology, Ohio State U. Medical Center, S305 Rhodes Hall, 450 W 10th Ave., Columbus, OH 43210. E-mail: gnuovomd@pol.net

In situ amplification permits the histological localization of low-copy DNA and RNA targets. However, in many instances it would be useful to know the specific phenotype of the target-containing cell or to ascertain the distribution of a different nucleic acid sequence in the same tissue section. This review describes a methodology that allows co-in situ localization of two nucleic acid targets or a DNA/RNA sequence and a protein in paraffin-embedded, formalin-fixed tissue. The key variable for detection of low-copy RNA targets by RT in situ PCR is optimal protease digestion to permit cDNA target-specific incorporation of the reporter nucleotide. This is achieved via inactivation of nonspecific DNA synthesis by overnight DNase digestion. The key variable for immunohistochemical localization of proteins is to determine the effect of protease digestion on the antigen-based signal intensity. Background for DNA targets by in situ hybridization or, for targets present in 1–10 copies per cell, PCR ISH is dependent primarily on probe concentration and the stringency of the post-hybridization wash. Radioactive 3H-labeled nucleotides permit an excellent distinction with colorimetric signals for co-localization, although two distinct chromogens can in many instances allow successful localization of two different targets.

(J Histochem Cytochem 49:1329–1339, 2001)

Key Words: in situ, PCR, RT in situ PCR, reverse transcriptase, immunohistochemistry


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
 BloodHome page
D. Fu and D. R. Richardson
Iron chelation and regulation of the cell cycle: 2 mechanisms of posttranscriptional regulation of the universal cyclin-dependent kinase inhibitor p21CIP1/WAF1 by iron depletion
Blood, July 15, 2007; 110(2): 752 - 761.
[Abstract] [Full Text] [PDF]

Home page
 GutHome page
J-Y Scoazec
Tissue and cell imaging in situ: potential for applications in pathology and endoscopy
Gut, June 1, 2003; 52(90004): iv1 - 6.
[Abstract] [Full Text] [PDF]

Home page
 J. Histochem. Cytochem.Home page
E. G. Tzortzaki, D. Glass, M. Yang, A. P. Evan, S. B. Bledsoe, P. J. Stambrook, A. Sahota, and J. A. Tischfield
Gender- and Age-dependent Changes in Kidney Androgen Protein mRNA Expression in a Knockout Mouse Model for Nephrolithiasis
J. Histochem. Cytochem., December 1, 2002; 50(12): 1663 - 1669.
[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 © 2001