HOMEPRODUCTSCOMPANYCONTACTFAQResearchDictionaryPharmaSign Up FREE or Login

ImmunoAT method: An initial assessment for the detection of abnormal isoforms of prion protein in formalin-fixed and paraffin-embedded tissues.

Abstract
The AT-tailing method is a labelling technique that utilises oligo(dA-dT)-dependent signal amplification. In this study, a new immunohistochemical application of the immunoAT method was developed. This method uses an oligo(dA-dT)-conjugated primary antibody (direct immunoAT method) or an oligo(dA-dT)-conjugated secondary antibody (indirect immunoAT method). Fifteen-base oligo(dA-dT)-conjugated antibodies (IgG-ATs) were prepared in advance by conjugating maleimide-activated oligo(dA-dT) to IgG via free sulfhydryl residues that had been introduced on the surface of IgG using Traut's reagent. Following the reaction with the target antigen and the IgG-AT, oligo(dA-dT) was elongated by DeltaTth DNA polymerase in the presence of dATP, dTTP and biotinylated dUTP, consequently labelling the antigen-antibody complex with a large amount of biotin. To initially evaluate the immunoAT method, the presence or absence of prion protein (PrP(sc)) was determined in formalin-fixed and paraffin-embedded sections of the medulla oblongata of cattle which had been under active surveillance for bovine spongiform encephalopathy. Sections were examined using direct and indirect immunoAT methods and the EnVision+ system (Dako) under conditions that were identical except for the differing IgG-AT and AT-tailing methods. PrP(sc) detection was consistent using all three methods. The clearest signals were obtained using the indirect immunoAT method, suggesting significant potential for this method.
AuthorsYuko Sato, Nozomi Shimonohara, Ken-Ichi Hanaki, Motoki Goto, Yoshio Yamakawa, Motohiro Horiuchi, Hidehiro Takahashi, Tetsutaro Sata, Noriko Nakajima
JournalJournal of virological methods (J Virol Methods) Vol. 165 Issue 2 Pg. 261-7 (May 2010) ISSN: 1879-0984 [Electronic] Netherlands
PMID20152861 (Publication Type: Evaluation Study, Journal Article, Research Support, Non-U.S. Gov't)
CopyrightCopyright 2010 Elsevier B.V. All rights reserved.
Chemical References
  • Antibodies
  • Oligodeoxyribonucleotides
  • PrPSc Proteins
  • oligo(dA-dT)
  • Formaldehyde
  • Poly dA-dT
Topics
  • Animals
  • Antibodies (metabolism)
  • Cattle
  • Encephalopathy, Bovine Spongiform (diagnosis)
  • Formaldehyde
  • Immunohistochemistry (methods)
  • In Situ Hybridization
  • Medulla Oblongata (chemistry, pathology)
  • Oligodeoxyribonucleotides (chemistry)
  • Paraffin Embedding
  • Poly dA-dT (chemistry)
  • PrPSc Proteins (immunology, isolation & purification)
  • Tissue Fixation

Join CureHunter, for free Research Interface BASIC access!

Take advantage of free CureHunter research engine access to explore the best drug and treatment options for any disease. Find out why thousands of doctors, pharma researchers and patient activists around the world use CureHunter every day.
Realize the full power of the drug-disease research graph!


Choose Username:
Email:
Password:
Verify Password:
Enter Code Shown: