PMID- 9792422
OWN - NLM
STAT- MEDLINE
DCOM- 19981211
LR  - 20191102
IS  - 0948-6143 (Print)
IS  - 0948-6143 (Linking)
VI  - 110
IP  - 4
DP  - 1998 Oct
TI  - Fluorescence in situ hybridization using horseradish peroxidase-labeled 
      oligodeoxynucleotides and tyramide signal amplification for sensitive DNA and 
      mRNA detection.
PG  - 431-7
AB  - We have used horseradish peroxidase-labeled 40 mer oligodeoxynucleotides 
      (HRP-ODNs) specific for the human cytomegalovirus immediate early gene (HCMV-IE) 
      and a novel dinitrophenol-tyramide signal amplification reagent (DNP-TSA plus) to 
      evaluate their utility in fluorescence in situ hybridization (FISH). For DNA 
      FISH, single or cocktails of HRP-ODNs were hybridized to metaphase chromosomes of 
      rat 9G cells which, as determined by DNA fiber FISH, carry an integrated tandem 
      repeat of 50-60 copies of the HCMV-IE gene. With one layer of DNP-TSA plus 
      deposition and subsequent detection with a fluorochrome-conjugated antibody, four 
      HRP-ODNs were needed to detect the HCMV-IE integration site. When employing two 
      TSA amplification rounds, one HRP-ODN was sufficient for obtaining a strong 
      signal of the integrated gene cluster, indicating that 50-60 HRP molecules can be 
      detected with ease. In addition to DNA FISH, we report here the first use of 
      HRP-ODN probes for mRNA detection by FISH. A single HRP-ODN and one DNP-TSA plus 
      step resulted in clear visualization of the HCMV-IE gene transcripts in rat 9G 
      cells induced for HCMV-IE expression by cycloheximide. Two TSA detection steps 
      enhanced signal intensities even further. Parallel experiments with 
      hapten-labeled ODN and cDNA probes and conventional detection methods illustrated 
      the superiority of the HRP-ODN/TSA approach in DNA and RNA FISH.
FAU - van de Corput, M P
AU  - van de Corput MP
AD  - Department of Molecular Cell Biology, Leiden University Medical Centre.
FAU - Dirks, R W
AU  - Dirks RW
FAU - van Gijlswijk, R P
AU  - van Gijlswijk RP
FAU - van de Rijke, F M
AU  - van de Rijke FM
FAU - Raap, A K
AU  - Raap AK
LA  - eng
PT  - Journal Article
PT  - Research Support, Non-U.S. Gov't
PL  - Germany
TA  - Histochem Cell Biol
JT  - Histochemistry and cell biology
JID - 9506663
RN  - 0 (Antigens, Viral)
RN  - 0 (DNA, Viral)
RN  - 0 (Dinitrophenols)
RN  - 0 (Immediate-Early Proteins)
RN  - 0 (Indicators and Reagents)
RN  - 0 (Oligonucleotide Probes)
RN  - 0 (RNA, Messenger)
RN  - 0 (RNA, Viral)
RN  - 0 (immediate-early proteins, cytomegalovirus)
RN  - EC 1.11.1.- (Horseradish Peroxidase)
SB  - IM
MH  - Animals
MH  - Antigens, Viral/genetics
MH  - Cell Line
MH  - Cloning, Molecular
MH  - Cytomegalovirus/*genetics
MH  - DNA, Viral/*analysis
MH  - Dinitrophenols
MH  - Genes, Immediate-Early
MH  - *Horseradish Peroxidase
MH  - Humans
MH  - Immediate-Early Proteins/genetics
MH  - In Situ Hybridization, Fluorescence/*methods
MH  - Indicators and Reagents
MH  - Metaphase
MH  - *Oligonucleotide Probes
MH  - RNA, Messenger/*analysis
MH  - RNA, Viral/analysis
MH  - Rats
MH  - Virus Integration
EDAT- 1998/10/29 00:00
MHDA- 1998/10/29 00:01
CRDT- 1998/10/29 00:00
PHST- 1998/10/29 00:00 [pubmed]
PHST- 1998/10/29 00:01 [medline]
PHST- 1998/10/29 00:00 [entrez]
AID - 10.1007/s004180050304 [doi]
PST - ppublish
SO  - Histochem Cell Biol. 1998 Oct;110(4):431-7. doi: 10.1007/s004180050304.