Rapid fluorescence detection of in situ hybridization with biotinylated bovine herpesvirus-1 DNA probes.
The use of biotinylated DNA hybridization probes for clinical detection of bovine herpesvirus-1 was investigated. Biotinylated DNA hybridization probes were prepared from bovine herpesvirus-1 DNA purified from infected cell cultures. The viral DNA was nick translated in the presence of biotin-dUTP with DNA polymerase to incorporate biotin into the newly synthesized strand. The probe was tested for specificity in in situ hybridization assays with bovine herpesvirus-1 DNA. Hybridization was detected using avidin-fluorescein single sandwich systems and an avidin-globulin with anti-globulin-fluorescein double sandwich system. Hybridization was detected by specific fluorescence of infected cells. Fluorescence was present only in bovine herpesvirus-1-infected cell culture and not in noninfected cell culture or cell cultures infected with several other viruses. The assay was performed in 6 hr.
Brock
K V
KV
Department of Pathobiology, University of Tennessee, College of Veterinary Medicine, Knoxville 37901.
Potgieter
L N
LN
D000818:Animals; D001360:Avidin / Q000031:analogs & derivatives; D001710:Biotin; D002417:Cattle; D002478:Cells, Cultured; D015342:DNA Probes; D004279:DNA, Viral / Q000235:genetics; D016650:Fluorescein-5-isothiocyanate / Q000031:analogs & derivatives*; D005452:Fluoresceins; D007242:Herpesvirus 1, Bovine / Q000235:genetics / Q000302:isolation & purification*; D008856:Microscopy, Fluorescence; D009693:Nucleic Acid Hybridization; D014176:Protein Biosynthesis
D015342:DNA Probes; D004279:DNA, Viral; D005452:Fluoresceins; C036963:fluorescein isothiocyante avidin; D001360:Avidin; D001710:Biotin; D016650:Fluorescein-5-isothiocyanate