Synthesis and purification of horseradish peroxidase-labeled oligonucleotides for tyramide-based fluorescence in situ hybridization.
A method is presented to conjugate horseradish peroxidase (HRP) to oligodeoxynucleotides for fluorescence in situ hybridization assays employing tyramide signal amplification (TSA). HRP is covalently bound to the oligonucleotide by thiol ether linkage and purified by high-performance liquid chromatography. With TSA detection, a single HRP-labeled oligonucleotide probe is sufficient for in situ detection of clustered DNA repeat sequences with a degree of repetition between 20 and 50.
van Gijlswijk
R P
RP
Department of Molecular Cell Biology, Leiden University Medical Center, The Netherlands.
van de Corput
M P
MP
Bezrookove
V
V
Wiegant
J
J
Tanke
H J
HJ
Raap
A K
AK
D002850:Chromatography, Gel; D002877:Chromosomes, Human / Q000737:chemistry; D004275:DNA, Ribosomal / Q000032:analysis; D005456:Fluorescent Dyes; D006735:Horseradish Peroxidase / Q000138:chemical synthesis*; D006801:Humans; D017404:In Situ Hybridization, Fluorescence / Q000379:methods*; D008297:Male; D015345:Oligonucleotide Probes / Q000737:chemistry; D009841:Oligonucleotides / Q000138:chemical synthesis*; D014443:Tyrosine / Q000031:analogs & derivatives*
D004275:DNA, Ribosomal; D005456:Fluorescent Dyes; D015345:Oligonucleotide Probes; D009841:Oligonucleotides; C105107:TSA-Direct GreenFISH; D014443:Tyrosine; D006735:Horseradish Peroxidase